Abstract
Here, the effects of copper stress (CS) on biomass accumulation, physiological parameters of oxidative stress and flavonoid production were investigated in Belamcanda chinensis calli. They were subjected to copper chloride treatments, and the corresponding responses were continuously monitored. Both concentration- and time-dependent physiological changes were uncovered using a multi-index evaluation. The changes in superoxide dismutase (SOD) and guaiacol peroxidase (GPX) activities could be divided into rising and descending stages, with the strongest activities occurring on d 28 and 35, respectively. Similar changes were observed in total flavonoid production, but the maximum yields occurred on different days at different copper chloride concentrations. During the experiment, the level of proline, which functioned in the early stage, declined and was replaced by other protective mechanisms, such as those provided by SOD and GPX, in the late stage. The SOD activity increased along with the copper concentration. However, the GPX activity decreased as the copper concentration increased. Proline’s trend was similar to that of GPX in the early stage but resembled that of SOD in the late stage. Notably, the contents and numbers of flavonoids increased as the copper concentration increased. When the concentration was 200 mg L−1, 11 kinds of flavonoids were generated at the milligram level, which indicates that this approach is potentially valuable for producing biologically active flavonoids. Finally, using a comprehensive analysis of the measured parameters, we concluded that plant cells resisted copper stress by activating multiple antioxidative defense mechanisms in a specific order and complementary manner.
Key message
Belamcanda chinensis has a remarkable ability to resist copper stress using enzymatic and non-enzymatic defenses, which are activated in a specific order and complementary manner.
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Abbreviations
- CS:
-
Copper stress
- DW:
-
Dry weight
- MS:
-
Murashige and Skoog
- FW:
-
Fresh weight
- MDA:
-
Malondialdehyde
- GPX:
-
Guaiacol peroxidase
- ROS:
-
Reactive oxygen species
- SOD:
-
Superoxide dismutase
- TFs:
-
Total flavonoids
References
Bartels D (2001) Targeting detoxification pathways: an efficient approach to obtain plants with multiple stress tolerance. Trends Plant Sci 6:284–286. https://doi.org/10.1016/S1360-1385(01)01983-5
Bates LS, Waldren RP, Teare ID (1973) Rapid determination of free proline for water-stress studies. Plant soil 39:205–207. https://doi.org/10.1007/BF00018060
Cao Y, Zhang Y, Ma C, Li H, Zhang J, Chen G (2018) Growth, physiological responses, and copper accumulation in seven willow species exposed to Cu-a hydroponic experiment. Environ Sci Pollut Res 25:19875–19886. https://doi.org/10.1007/s11356-018-2106-z
Chen CT, Chen TH, Lo KF, Chiu CY (2004) Effects of proline on copper transport in rice seedlings under excess copper stress. Plant Sci 166:103–111. https://doi.org/10.1016/j.plantsci.2003.08.015
Chen Y, Wu CM, Dai RJ, Li L, Yu YH, Li Y, Meng WW, Zhang L, Zhang YQ, Deng YL (2011) Combination of HPLC chromatogram and hypoglycemic effect identifies isoflavones as the principal active fraction of Belamcanda chinensis leaf extract in diabetes treatment. J Chromatogr B 879:371–378. https://doi.org/10.1016/j.jchromb.2010.12.022
Chen Y, Luo YY, Qiu NF, Hu F, Sheng LL, Wang RQ, Cao FL (2015) Ce3+ induces flavonoids accumulation by regulation of pigments, ions, chlorophyll fluorescence and antioxidant enzymes in suspension cells of Ginkgo biloba L. Plant Cell Tiss Organ Cult 123:283–296. https://doi.org/10.1007/s11240-015-0831-2
Chinese Pharmacopoeia Committee (2015) Pharmacopoeia of the People’s Republic of China. Chinese Medical Science and Technology Press, Beijing, p 285
Drążkiewicz M, Skórzyńska-Polit E, Krupa Z (2004) Copper-induced oxidative stress and antioxidant defence in Arabidopsis thaliana. Biometals 17:379–387. https://doi.org/10.1023/B:BIOM.0000029417.18154.22
Faisal M, Anis M (2009) Changes in photosynthetic activity, pigment composition, electrolyte leakage, lipid peroxidation, and antioxidant enzymes during ex vitro establishment of micropropagated Rauvolfia tetraphylla plantlets. Plant Cell Tissue Organ Cult 99:125–132. https://doi.org/10.1007/s11240-009-9584-0
Ghoulam C, Foursy A, Fares K (2002) Effects of salt stress on growth, inorganic ions and proline accumulation in relation to osmotic adjustment in five sugar beet cultivars. Environ Exp Bot 47:39–50. https://doi.org/10.1016/S0098-8472(01)00109-5
Gichner T, Patková Z, Száková J, Demnerová K (2006) Toxicity and DNA damage in tobacco.nd potato plants growing on soil polluted with heavy metals. Ecotox Environ Safe 65:420–426. https://doi.org/10.1016/j.ecoenv.2005.08.006
Hu H, Jin Q, Kavan P (2014) A study of heavy metal pollution in China: current status, pollution-control policies and countermeasures. Sustainability 6:5820–5838. https://doi.org/10.3390/su6095820
Jabeen R, Ahmad A, Iqbal M (2009) Phytoremediation of heavy metals: physiological and molecular mechanisms. Bot Rev 75:339–364. https://doi.org/10.1007/s12229-009-9036-x
Kalir A, Ornri G, Poljakoff-Mayber A (1984) Peroxidase and catalase activity in leaves of Halimione portulacoides exposed to salinity. Physiol Plant 62:238–244. https://doi.org/10.1111/j.1399-3054.1984.tb00377.x
Khatun S, Ali MB, Hahn EJ, Paek KY (2008) Copper toxicity in Withania somnifera: growth and antioxidant enzymes responses of in vitro grown plants. Environ Exp Bot 64:279–285. https://doi.org/10.1016/j.envexpbot.2008.02.004
Ku HM, Tan CW, Su YS, Chiu CY, Chen CT, Jan FJ (2012) The effect of water deficit and excess copper on proline metabolism in Nicotiana benthamiana. Biol Plant 56:337–343. https://doi.org/10.1007/s10535-012-0095-1
Li LM, Wang RG (2012) Effects of NaCl stress on physiological and biochemical characteristics of Belamcanda chinensis and zirs germanic two ground cover plants. Chin Agric Sci Bull 28:181–184
Liu ML, Li XR, Liu YB, Cao B (2013) Regulation of flavanone 3-hydroxylase gene involved in the flavonoid biosynthesis pathway in response to UV-B radiation and drought stress in the desert plant, Reaumuria soongorica. Plant Physiol Biochem 73:161–167. https://doi.org/10.1016/j.plaphy.2013.09.016
Lokhande VH, Nikam TD, Patade VY, Ahire ML, Suprasanna P (2011) Effects of optimal and supra-optimal salinity stress on antioxidative defence, osmolytes and in vitro growth responses in Sesuvium portulacastrum L. Plant Cell Tissue Organ Cult 104:41–49. https://doi.org/10.1007/s11240-010-9802-9
Lombardi L, Sebastiani L (2005) Copper toxicity in Prunuscerasifera: growth and antioxidant enzymes responses of in vitro grown plants. Plant Sci 168:797–802. https://doi.org/10.1016/j.plantsci.2004.10.012
Lukatkin A, Egorova I, Michailova I, Malec P, Strzałka K (2014) Effect of copper on pro- and antioxidative reactions in radish (Raphanus sativus L.) in vitro and in vivo. J Trace Elem Med Bio 28:80–86. https://doi.org/10.1016/j.plantsci.2004.10.012
Maheshwari R, Dubey RS (2007) Nickel toxicity inhibits ribonuclease and protease activities in rice seedlings: protective effects of proline. Plant Growth Regul 51:231–243. https://doi.org/10.1007/s10725-006-9163-x
Moghaieb REA, Saneoka H, Fujita K (2004) Effect of salinity on osmotic adjustment, glycinebetaine accumulation and the betaine aldehyde dehydrogenase gene expression in two halophytic plants, Salicornia europaea and Suaeda maritima. Plant Sci 166:1345–1349. https://doi.org/10.1016/j.plantsci.2004.01.016
Mostofa MG, Hossain MA, Fujita M, Tran LSP (2015) Physiological and biochemical mechanisms associated with trehalose-induced copper-stress tolerance in rice. Sci Rep. https://doi.org/10.1038/srep11433
Mourato M, Reis R, Martins MLL (2012) Characterization of plant antioxidative system in response to abiotic stresses: a focus on heavy metal toxicity. Advances in selected plant physiology aspects, Croatia, pp 23–44
Posmyk MM, Kontek R, Janas KM (2009) Antioxidant enzymes activity and phenolic compounds content in red cabbage seedlings exposed to copper stress. Ecotox Environ Safe 72:596–602. https://doi.org/10.1016/j.ecoenv.2008.04.024
Procházková D, Boušová I, Wilhelmová N (2011) Antioxidant and prooxidant properties of flavonoids. Fitoterapia 82:513–523. https://doi.org/10.1016/j.fitote.2011.01.018
Qin MJ, Ji WL, Liu J, Yu G (2003) Scavenging effects on radicals of isoflavones from rhizome of Belamcandae chinensis. Chin Tradit Herb Drugs 34:640–641
Rai PK, Lee SS, Zhang M, Tsang YF, Kim KH (2019) Heavy metals in food crops: Health risks, fate, mechanisms, and management. Environ Int 125:365–385. https://doi.org/10.1016/j.envint.2019.01.067
Rejeb KB, Abdelly C, Savouré A (2014) How reactive oxygen species and proline face stress together. Plant Physiol Biochem 80:278–284. https://doi.org/10.1016/j.plaphy.2014.04.007
Seidlova-Wuttke D, Hesse O, Jarry H, Rimoldi G, Thelen P, Christoffel V, Wuttke W (2004) Belamcanda chinensis and the thereof purified tectorigenin have selective estrogen receptor modulator activities. Phytomedicine 11:392–403. https://doi.org/10.1016/j.phymed.2004.01.003
Singh S, Parihar P, Singh R, Singh VP, Prasad SM (2016) Heavy metal tolerance in plants: role of transcriptomics, proteomics, metabolomics, and lonomics. Front Plant Sci 6:1143. https://doi.org/10.3389/fpls.2015.01143
Singh S, Singh S, Ramachandran V, Eapen S (2010) Copper tolerance and response of antioxidative enzymes in axenically grown Brassica juncea plants. Ecotox Environ Safe 73:1975–1981. https://doi.org/10.1016/j.ecoenv.2010.08.020
Smirnoff N, Cumbes QJ (1989) Hydroxyl radical scavenging activity of compatible solutes. Phytochemistry 28:1057–1060. https://doi.org/10.1016/0031-9422(89)80182-7
Szabados L, Savouré A (2010) Proline: a multifunctional amino acid. Trends Plant Sci 15:89–97. https://doi.org/10.1016/j.tplants.2009.11.009
Tebayashi S, Ishihara A, Iwamura H (2001) Elicitor- induced changes in isoflavonoid metabolism in red clover roots. J Exp Bot 52:681–689. https://doi.org/10.1093/jexbot/52.357.681
Tewari RK, Kumar P, Sharma PN (2006) Antioxidant responses to enhanced generation of superoxide anion radical and hydrogen peroxide in the copper-stressed mulberry plants. Planta 223:1145–1153. https://doi.org/10.1007/s00425-005-0160-5
Thounaojam TC, Panda P, Mazumdar P, Kumar D, Sharma GD, Sahoo L, Sanjib P (2012) Excess copper induced oxidative stress and response of antioxidants in rice. Plant Physiol Bioch 53:33–39. https://doi.org/10.1016/j.plaphy.2012.01.006
Tian GZ, Li HF, Qiu WF (2001) Advances on research of plant peroxidases. J Wuhan Bot Res 19:332–344
Trettel JR, Gazim ZC, GonÇalves JE, Stracieri J, Magalhães HM (2019) Effects of copper sulphate (CuSO4) elicitation on the chemical constitution of volatile compounds and the in vitro development of Basil. Sci Hortic 234:19–26. https://doi.org/10.1016/j.scienta.2018.01.062
Treutter D (2006) Significance of flavonoids in plant resistance: a review. Environ Chem Lett 4:147–157. https://doi.org/10.1007/s10311-006-0068-8
Vaidyanathan H, Sivakumar P, Chakrabarty R, Thomas G (2003) Scavenging of reactive oxygen species in NaCl-stressed rice (Oryza sativa L.)-differential response in salt-tolerant and sensitive varieties. Plant Sci 165:1411–1418. https://doi.org/10.1016/j.plantsci.2003.08.005
Wang LH, An MJ, Huang WD, Zhan JC (2019) Melatonin and phenolics biosynthesis-related genes in Vitis vinifera cell suspension cultures are regulated by temperature and copper stress. Plant Cell Tiss Organ Cult 138:475–488. https://doi.org/10.1007/s11240-019-01643-1
Xie GY, Zhu Y, Shu P, Qin XY, Wu G, Wang Q, Qin MJ (2014) Phenolic metabolite profiles and antioxidants assay of three Iridaceae medicinal plants for traditional Chinese medicine “She-gan” by on-line HPLC–DAD coupled with chemiluminescence (CL) and ESI-Q-TOF-MS/MS. J Pharmaceut Biomed 98:40–51. https://doi.org/10.1016/j.jpba.2014.05.008
Xu MP, He P, Duan CX, Yang M (2014) Study on relieving effects of exogenous SNP, Spd on Belamcanda chinensis under salt-alkalline stress. Chin J Chin Mater Med 39:4553–4558
Xu XY, Xie GS, He L, Zhang JJ, Xu XL, Qian R, Liang GH, Liu JH (2013) Differences in oxidative stress, antioxidant systems, and microscopic analysis between regenerating callus-derived protoplasts and recalcitrant leaf mesophyll-derived protoplasts of Citrus reticulata Blanco. Plant Cell Tissue Organ Cult 114:161–169. https://doi.org/10.1007/s11240-013-0312-4
Xu XY, Shi GX, Wang J, Kang YN (2011) Copper-induced oxidative stress in Alternanthera philoxeroides callus. Plant Cell Tissue Organ Cult 106:243–251. https://doi.org/10.1007/s11240-010-9914-2
Yang M, He P, Duan CX, Xu MP (2015) Effects of exogenous SNP and Spd on the activity of antioxidant enzyme system in Belamacanda chinensis (L.) DC. seedlings under saline-alkali stress. J Southw Univ 37:13–19
Yruela I (2009) Copper in plants: acquisition, transport and interactions. Funct Plant Biol 36:409–430. https://doi.org/10.1071/FP08288
Zhang LP, Mehta SK, Liu ZP, Yang ZM (2008) Copper-induced proline synthesis is associated with nitric oxide generation in Chlamydomonas reinhardtii. Plant Cell Physiol 49:411–419. https://doi.org/10.1093/pcp/pcn017
Zhang YY, Wang Q, Qi LW, Qin XY, Qin MJ (2011) Characterization and determination of the major constituents in Belamcandae Rhizoma by HPLC–DAD–ESI-MS n. J Pharmaceut Biomed 56:304–314. https://doi.org/10.1016/j.jpba.2011.05.040
Zhu Y, Pu BQ, Xie GY, Tian M, Xu FY, Qin MJ (2014) Dynamic changes of flavonoids contents in the different parts of rhizome of Belamcanda chinensis during the thermal drying Process. Molecules 19:10440–10454. https://doi.org/10.3390/molecules190710440
Acknowledgements
This work was supported by the National Natural Science Foundation of China (Grant No. 81872958 and 81373918), the National Major Scientific and Technological Special Project for “Significant New Drugs Development” during the Twelfth Five-year Plan Period (Grant No. 2012ZX09304006) and the Fundamental Research Funds for the Central Universities (JKQ2011038 and PT2014ZY0086).
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YZ and MJQ designed research; YZ, YHC and XZ performed research and all authors analyzed the data; YZ, YHC and MJQ wrote the paper. All authors read and approved the final manuscript.
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Zhu, Y., Chen, Y., Zhang, X. et al. Copper stress-induced changes in biomass accumulation, antioxidant activity and flavonoid contents in Belamcanda chinensis calli. Plant Cell Tiss Organ Cult 142, 299–311 (2020). https://doi.org/10.1007/s11240-020-01863-w
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DOI: https://doi.org/10.1007/s11240-020-01863-w