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Large-scale isolation of scopoletin from Nicotiana tabacum

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Abstract

Nicotiana tabacum leaves are mainly used in the production of cigarettes and cigars all over the world, and a large amount of waste materials will be produced in the process. High-value scopoletin can be extracted and separated from waste materials, but there are some problems, such as low purity of products and inability of large-scale production. In this study, we established an efficient and stable method for producing high-purity scopoletin from waste tobacco by containing acetone reflux extracting with dynamic axial compression-high performance liquid chromatography (DAC-HPLC) separating. In particular, the extraction system makes the scopoletin fully extracted through continuous reflux, the DAC-HPLC system makes the high-purity scopoletin repeatedly obtained, and the DAC-HPLC system was first used for scopoletin. Finally, related to the waste tobacco power, the yield of scopoletin is 1.9%, and its HPLC purity is 99.94%. Additionally, a preparation method for the colorless needle crystal of scopoletin was established. Meanwhile, it has a strong antioxidant activity by testing the scavenging activity of the DPPH free radical and the Fe3+ reducing power. And its spatial structure was identified by an X-ray diffractometer, which laid a good foundation for in-depth study of scopoletin.

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All data generated or analyzed during this study are included in this manuscript and its supplementary information files.

Abbreviations

DAC-HPLC:

Dynamic axial compression-high performance liquid chromatography

C18:

Octadecylsilane (C18H40Si)

DPPH:

1,1-Diphenyl-2-picrylhydrazyl

TA:

Tobacco

SE:

Scopoletin extraction

Ve:

Vitamin E

References

  1. Andersen RA, Vaughn TH (1972) Short communication micro determination of scopoletin in Nicotiana tabacum. Phytochemistry 11:2593–2595

    Article  Google Scholar 

  2. Lerat S, Babana AH, El OM, Daayf F, Beaudoin N, Bouarab K, Beaulieu C et al (2009) Streptomyces scabiei and its toxin thaxtomin A induce scopoletin biosynthesis in tobacco and Arabidopsis thaliana. Plant Cell Rep 28:1895–1903

    Article  Google Scholar 

  3. Zhao HN, Cai K, Qin J et al (2017) Variance analysis of main metabolites in tobacco leaves before and after curing. Tob Sci Technol 50:61–67

    Google Scholar 

  4. Parama D, Girisa S, Khatoon E et al (2022) An overview of the pharmacological activities of scopoletin against different chronic diseases. Pharmacol Res 179:106202

    Article  Google Scholar 

  5. Agati G, Brunetti C, Tuccio L et al (2022) Retrieving the in vivo scopoletin fluorescence excitation band allows the non-invasive investigation of the plant-pathogen early events in tobacco leaves. Front Microbiol 13:889878

    Article  Google Scholar 

  6. Siwinska J, Siatkowska K, Olry A et al (2018) Scopoletin 8-hydroxylase: a novel enzyme involved in coumarin biosynthesis and iron-deficiency responses in Arabidopsis. J Exp Bot 69(7):1735–1748

    Article  Google Scholar 

  7. Tsai HH, Rodríguez-Celma J, Lan P et al (2018) Scopoletin 8-hydroxylase-mediated fraxetin production is crucial for iron mobilization. Plant Physiol 177(1):194–207. https://doi.org/10.1104/pp.18.00178

    Article  Google Scholar 

  8. Döll S, Kuhlmann M, Rutten T et al (2018) Accumulation of the coumarin scopolin under abiotic stress conditions is mediated by the Arabidopsis thaliana THO/TREX complex. Plant J 93(3):431–444

    Article  Google Scholar 

  9. Sun H, Song N, Ma L et al (2017) Ethylene signalling is essential for the resistance of Nicotiana attenuata against Alternaria alternata and phytoalexin scopoletin biosynthesis. Plant Pathol 66(2):277–284

    Article  Google Scholar 

  10. Choi RY, Ham JR, Lee HI et al (2017) Scopoletin supplementation ameliorates steatosis and inflammation in diabetic mice. Phytother Res 31(11):1795–1804

    Article  Google Scholar 

  11. Zhao YX, Wang M, Dong WT et al (2019) Pharmacokinetics, bioavailability and metabolism of scopoletin in dog by ultra-high-performance liquid chromatography combined with linear ion trap-Orbitrap tandem mass spectrometry. Biomed Chromatogr 33(3):e4436

    Article  Google Scholar 

  12. Fritig B, Hirth L, Ourisson G (1970) Biosynthesis of the Coumarins: Scopoletin formation in tobacco tissue cultures. Phytochemistry 9:1963–1975

    Article  Google Scholar 

  13. Sun H, Wang L, Zhang B (2014) Scopoletin is a phytoalexin against Alternaria alternata in wild tobacco dependent on jasmonate signalling. J Exp Bot 65:4305–4315

    Article  Google Scholar 

  14. Fourcroy P, Sisó-Terraza P, Sudre D et al (2014) Involvement of the ABCG37 transporter in secretion of scopoletin and derivatives by Arabidopsis roots in response to iron deficiency. New Phytol 201(1):155–167

    Article  Google Scholar 

  15. Stringlis IA, Yu K, Feussner K et al (2018) MYB72-dependent coumarin exudation shapes root microbiome assembly to promote plant health. Proc Natl Acad Sci USA 115(22):E5213–E5222

    Article  Google Scholar 

  16. Hoengenaert L, Wouters M, Kim H et al (2022) Overexpression of the scopoletin biosynthetic pathway enhances lignocellulosic biomass processing. Sci adv 8(28):eabo738

    Article  Google Scholar 

  17. Gao SN, Li S, Zhang LN et al (2012) Isolation and purification of scopoletin from Erycibe obtusifolia by high-speed counter-current chromatography. China Med Her 9(26):122–123

    Google Scholar 

  18. Sun JY, Zhou YQ, Li HY et al (2003) Determination of scopoletin in Cortex Mori from different sources. Chin Herb Med 03:77–79

    Google Scholar 

  19. Geng ZL, Zhang J, Yang MM et al (2013) Study on the chemical constituents of the tobacco leaves from Guizhou Province. J Yunnan Minzu University (Soc Sci) 22(2):79–81

    Google Scholar 

  20. Li L (2010) Study on the chemical composition of the fruit of Acanthopanax senticosus. Liaoning University of Traditional Chinese Medicine

    Google Scholar 

  21. Liu C, Hu Y, Li N et al (2009) Isolation of scopoletin from Physochlaine infundibularis Kuang by high-speed counter-current chromatography. Chem Res 20(02):101–103

    Google Scholar 

  22. Yang L, Wang MY, Zhang D et al (2006) Determination of scopoletin in Qinghao by HPLC. Chin J Exp Prescriptions 10:10–11

    Google Scholar 

  23. Sun YL, Liu WL, Liu YN et al (2018) Extraction technology of scopoletin from Rubus corchorifolius. Guizhou Agric Sci 46(05):120–122

    Google Scholar 

  24. Jokić S, Rajić M, Bilić B et al (2016) Supercritical extraction of scopoletin from Helichrysum italicum (Roth) G. Don flowers. Phytochem Anal 27(5):290–295

    Article  Google Scholar 

  25. Chen X, Zhu P, Liu B et al (2018) Simultaneous determination of fourteen compounds of Hedyotis diffusa Willd extract in rats by UHPLC-MS/MS method: application to pharmacokinetics and tissue distribution study. J Pharm Biomed Anal 159:490–512

    Article  Google Scholar 

  26. Sun HR, Yao H, Shi GF (2010) Study on chemical constituents from waste tobacco leavesII. Anhui Agric Sci 38:8394–8409

    Google Scholar 

  27. Kazuki T, Ayumi K, Takashi W et al (2019) Chemical constituents from the aerial parts of impatiens hypophylla Makino var. hypophylla. Biochem Syst Ecol 83:10–12

    Article  Google Scholar 

  28. Donald GC, Robert VB (1962) New syntheses in the coumarin series. J Org Chem 27:3083–3085

    Article  Google Scholar 

  29. Lang’at TC, Song TT, Hu J et al (2003) A simple synthesis of 7,4’-dihydroxy-6-methoxyisoflavone, glycitein, the third soybean isoflavone. J Nat Prod 66:149–151

    Article  Google Scholar 

  30. Fang Z, He GL, He L (2007) Microwave-assisted synthesis of scopolamine. West China Pharm J 22:302–303

    Google Scholar 

  31. He BT, Liu ZH, Li BZ et al (2022Aug 2) Advances in biosynthesis of scopoletin. Microb Cell Fact 21(1):152

    Article  Google Scholar 

  32. Rajniak J, Giehl R, Chang E et al (2018) Biosynthesis of redox-active metabolites in response to iron deficiency in plants. Nat Chem Biol 14(5):442–450

    Article  Google Scholar 

  33. Kai K, Mizutani M, Kawamura N (2008) Scopoletin is biosynthesized via ortho-hydroxylation of feruloyl CoA by A 2-oxoglutarate-dependent dioxygenase in Arabidopsis thaliana. Plant J 55(6):989–999

    Article  Google Scholar 

  34. Bayoumi SA, Rowan MG, Blagbrough IS et al (2008) Biosynthesis of scopoletin and scopolin in cassava roots during post-harvest physiological deterioration: the E-Z-isomerisation stage. Phytochemistry 69(17):2928–2936

    Article  Google Scholar 

  35. Yao R, Zhao Y, Liu T et al (2017) Identification and functional characterization of a P-coumaroyl CoA 2’-hydroxylase involved in the biosynthesis of coumarin skeleton from Peucedanum praeruptorum Dunn. Plant Mol Biol 95(1–2):199–213

    Article  Google Scholar 

  36. Geng ZL, Zhang J, Yang MG et al (2013) Study on the chemical composition of tobacco leaves in Guizhou (I). J Yunnan Univ National (Nat Sci Ed) 22:79–81

    Google Scholar 

  37. Chen Y, Wu Y, Li S et al (2020) Large-scale isolation and antitumor mechanism evaluation of compounds from the traditional Chinese medicine Cordycepsmilitaris. Eur J Med Chem 212:113142

    Article  Google Scholar 

  38. Gao H, Huang L, Ding F et al (2018) Simultaneous purification of dihydrotanshinone, tanshinone I, cryptotanshinone, and tanshinone IIA from Salvia miltiorrhiza and their anti-inflammatory activities investigation. Sci Rep 8(1):8460

    Article  Google Scholar 

  39. Dang J, Cui Y, Pei J et al (2018) Efficient separation of four antibacterial diterpenes from the roots of Salvia prattii using non-aqueous hydrophilic solid-phase extraction followed by preparative high-performance liquid chromatography. Molecules 23(3):623

    Article  Google Scholar 

  40. Cong J (2013) Separation and purification of superoxide dismutase (SOD) in wheat seedlings and its antioxidant activity. Sichuan Agricultural University

    Google Scholar 

  41. Wang YL (2018) Preparation of phosphatized Bletilla striata polysaccharide and its application in cosmetics. Shanghai University of Applied Sciences

    Google Scholar 

  42. Adfa M, Yoshimura T, Komura K et al (2010) Antitermite activities of coumarin derivatives and scopoletin from Protium javanicum Burm. f. J Chem Ecol 36:720–726

    Article  Google Scholar 

  43. Beh HK, Ismail Z, Asmawi MZ et al (2010) 7-Hy-droxy-6-meth-oxy-2H-chromen-2-one. Acta Crystallogr Sect E Struct Rep Online 66:o2138

    Article  Google Scholar 

  44. Banožić M, Banjari I, Jakovljević M et al (2019) Optimization of ultrasound-assisted extraction of some bioactive compounds from tobacco waste. Molecules 24(8):1611

    Article  Google Scholar 

  45. Laszlo C, Kaminski K, Guan H et al (2022) Fractionation and extraction optimization of potentially valuable compounds and their profiling in six varieties of two Nicotiana species. Molecules 27(22):8105

    Article  Google Scholar 

  46. Lemos ASO, Florêncio JR, Pinto NCC et al (2020) Antifungal activity of the natural coumarin scopoletin against planktonic cells and biofilms from a multidrug-resistant Candida tropicalis strain. Front Microbiol 11:1525

    Article  Google Scholar 

  47. Siwinska J, Kadzinski L, Banasiuk R et al (2014) Dentification of QTLs affecting scopolin and scopoletin biosynthesis in Arabidopsis thaliana. BMC Plant Biol 14:280

    Article  Google Scholar 

  48. Sun X, Zhou D, Kandavelu P et al (2015) Structural insights into substrate specificity of feruloyl-CoA 6’-hydroxylase from Arabidopsis thaliana. Sci Rep 5:10355

    Article  Google Scholar 

  49. Li J and Wu J (2016) Scopolin, a glycoside form of the phytoalexin scopoletin, is likely involved in the resistance of Nicotiana attenuata against Alternaria alternata. J Plant Pathol 98(3)

  50. Chen Y, Wu Y, Liu L et al (2019) Study of the whole genome, methylome and transcriptome of Cordycepsmlitaris. Sci Rep 9(1):898

    Article  Google Scholar 

  51. Siwinska J, Siatkowska K, Olry A (2018) Scopoletin 8-hydroxylase: a novel enzyme involved in coumarin biosynthesis and iron-deficiency responses in Arabidopsis. J Exp Bot 69:1735–1748

    Article  Google Scholar 

  52. Tsai HH, Rodríguez CJ, Lan P et al (2018) Scopoletin 8-hydroxylase-mediated fraxetin production is crucial for iron mobilization. Plant Physiol 177(1):194–207

    Article  Google Scholar 

  53. Sun X, Zhou D, Kandavelu P (2015) Structural insights into substrate specificity of feruloyl-CoA 6’-hydroxylase from Arabidopsis thaliana. Sci Rep 5:10355

    Article  Google Scholar 

  54. Hu S, Feng J, Wang M et al (2022) Nrf1 is an indispensable redox-determining factor for mitochondrial homeostasis by integrating multi-hierarchical regulatory networks. Redox Biol 57:102

    Article  Google Scholar 

  55. Zhu YP, Zheng Z, Hu S et al (2019) Unification of opposites between two antioxidant transcription factors Nrf1 and Nrf2 in mediating distinct cellular responses to the endoplasmic reticulum stressor tunicamycin. Antioxidants 9(1):4

    Article  Google Scholar 

  56. Ma XF, Zhang YY, Guo FY et al (2020) Molecular characterization of a voltage-gated calcium channel and its potential role in the acaricidal action of scopoletin against Tetranychus cinnabarinus. Pestic Biochem Physiol 168:104618

    Article  Google Scholar 

  57. Wang Y, Zhang K, Li T et al (2021) Macrophage membrane functionalized biomimetic nanoparticles for targeted anti-atherosclerosis applications. Theranostics 11(1):164–180

    Article  Google Scholar 

  58. Narasimhan KKS, Jayakumar D, Velusamy P et al (2019) Morinda citrifolia and its active principle scopoletin mitigate protein aggregation and neuronal apoptosis through augmenting the DJ-1/Nrf2/ARE signaling pathway. Oxid Med Cell Longev 2019:2761041

    Article  Google Scholar 

  59. Rong N, Yang R, Ibrahim IAA et al (2022) Cardioprotective role of scopoletin on isoproterenol-induced myocardial infarction in rats. Appl Biochem Biotechnol. https://doi.org/10.1007/s12010-022-04123-z

  60. Wang X, Wan M, Zhang L et al (2022) ALA_PDT promotes ferroptosis-like death of Mycobacterium abscessus and antibiotic sterilization via oxidative stress. Antioxidants 11(3):546

    Article  Google Scholar 

  61. Huang X, Nisar MF, Wang M et al (2020) UV-responsive AKBA@ZnO nanoparticles potential for PMLE protection and therapy. Mater Sci Eng C Mater Biol Appl 107:110254

    Article  Google Scholar 

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Acknowledgements

We would like to thank the China National Tobacco Corporation’s Guizhou Provincial Company for providing the tobacco raw materials, and Ling Hui (Guangzhou) Biotech Co. Ltd for promoting the wide application of scopoletin in this study. J.W. Bartsch, Marburg, Germany for useful discussions and editing.

Funding

This work were supported by the National Science and Technology Major Project, China (Grant Number: 2012ZX09401009), the Chongqing Municipal Education Commission, China (Grant Number: KYYJ202001), the Fundamental Research Funds for Central Universities, China (Grant Number: 2022CDJYGRH-005&2022CDJYGRH-016), and the Science and Technology Foundation, China (Grant Number: QKHZC [2017]2960, QKHZC [2018]2822).

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Authors and Affiliations

  1. Key Laboratory for Biorheological Science and Technology of Ministry of Education, State and Local Joint Engineering Laboratory for Vascular Implants, Bioengineering College of Chongqing University, Chongqing, 400030, China

    Yujiao Chen, Yidan Chen, Li Zhong & Guixue Wang

  2. Guizhou Aerospace Intelligent Agriculture Co. Ltd., Guizhou, China

    Yujiao Chen, Xiaowei Su, Jingwen Zeng & Zhengbo Liu

  3. Guizhou Guian Precision Medicine Co. Ltd., Gui’an, Guizhou, China

    Yujiao Chen, Wuhua Zhang, Shishan Zhang, Taijun Zhao & Jun Cao

  4. Guizhou Yangyanshi Technology Co., Ltd., Guizhou, China

    Min Chen

  5. Guizhou Tobacco Science Research Institute, Guizhou, China

    Xiankun Su

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  1. Yujiao Chen
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  2. Min Chen
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Contributions

Y.-J. C. designed the study and wrote the manuscript. M. C., W.-H. Z., and S.-S. Z. performed experiments. X.-K. S., T.-J. Z., X.-W. S., J.-W.Z., and Y.-D. C. contributed to some experiments. G.-X. W., L. Z., J. C., and Z.-B. L. supervised the study. All authors read and approved the final manuscript.

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Correspondence to Yujiao Chen, Zhengbo Liu, Li Zhong or Guixue Wang.

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Chen, Y., Chen, M., Zhang, W. et al. Large-scale isolation of scopoletin from Nicotiana tabacum. Biomass Conv. Bioref. (2023). https://doi.org/10.1007/s13399-023-03778-w

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