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Enrichment and purification of red pigments from defective mulberry fruits using biotransformation in a liquid-liquid-solid three-phase system

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Abstract

A large number of defective mulberries are discarded each year because mulberries are easy to break. The red pigments from defective mulberries are recognized as the sustainable sources of anthocyanins extracted from nature. Cyanidin-3-O-rutinoside and cyanidin-3-O-glucoside are the main components of mulberry red pigments, accounting for 50% and 40% of the total, respectively. Cyanidin-3-O-glucoside exhibits anticancer, hypoglycemic, and liver and visceral protection properties. Cyanidin-3-O-glucoside can be prepared by enzymatically hydrolyzing the rhamnosidase bond of cyanidin-3-O-rutinoside. To obtain mulberry red pigment with a high purity of cyanidin-3-O-glucoside, immobilized α-L-rhamnosidase was added to the aqueous two-phase system to construct a liquid-liquid-solid three-phase enzyme catalytic system. After optimization, the three-phase system was composed of 27.12% (w/w) ethanol, 18.10% (w/w) ammonium sulfate, 15% (w/w) mulberry juice, 4.24% (w/w) immobilized α-L-rhamnosidase, and 35.54% (w/w) pure water. The three-phase system was employed to enrich and purify cyanidin-3-O-glucoside at pH 5 and 45 °C for 1 h. The purity of cyanidin-3-O-glucoside was increased from 40 to 82.42% with cyanidin-3-O-rutinoside conversion of 60.68%. The immobilized α-L-rhamnosidase could be reused seven times, maintaining a relative activity of over 50%. Overall, the developed system provided an efficient and simple approach for high purity mulberry red pigment production and recycling in the field of sustainable agriculture.

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References

  • Al-Musawi TJ, Brouers F, Zarrabi M (2017) Kinetic modeling of antibiotic adsorption onto different nanomaterials using the Brouers-Sotolongo fractal equation. Environ Sci Pollut Res 24:4048–4057

    Article  CAS  Google Scholar 

  • Antón-Millán N, Garcia-Tojal J, Marty-Roda M, Garroni S, Cuesta-López S, Tamayo-Ramos JA (2018) Influence of three commercial graphene derivatives on the catalytic properties of a Lactobacillus plantarum α-L-rhamnosidase when used as immobilization matrices. ACS Appl Mater Interfaces 10:18170–18182

    Article  Google Scholar 

  • Arca-Ramos A, Ammann EM, Gasser CA, Nastold P, Eibes G, Feijoo G, Lema JM, Moreira MT, Corvini PFX (2016) Assessing the use of nanoimmobilized laccases to remove micropollutants from wastewater. Environ Sci Pollut Res 23:3217–3228

    Article  CAS  Google Scholar 

  • Bjoroy O, Rayyan SF, Torgils, Kalberg K, Andersen OM (2009) C-glycosylanthocyanidins synthesized from C-glycosylflavones. Phytochemistry 70:278–287

    Article  Google Scholar 

  • Chen ZQ, Wang C, Pan YX, Gao XD, Chen HX (2017) Hypoglycemic and hypolipidemic effects of anthocyanins extract from black soybean seed coat in high fat diet and streptozotocin-induced diabetic mice. Food Funct 9:426–439

    Article  Google Scholar 

  • Cheng J, Lou WY, Zong MH (2014) Biocatalytic asymmetric oxidation of racemic 1-(4-methoxyphenyl) ethanol using immobilized Acetobacter sp.CCTCC M209061 cells in organic solvent-containing biphasic system. Chem J Chin Univ 35:1529–1535

    CAS  Google Scholar 

  • Cruz L, Benohoud M, Rayner CM, Mateus N, Freitas VD, Blackburn RS (2018) Selective enzymatic lipophilization of anthocyanin glucosides from blackcurrant (Ribes nigrum L.) skin extract and characterization of esterified anthocyanins. Food Chem 266:415–419

    Article  CAS  Google Scholar 

  • Eijsink VG, Bjørk A, Gåseidnes S, Sirevåg R, Synstad B, Van B, Vriend G (2004) Rational engineering of enzyme stability. J Biotechnol 113:105–120

    Article  CAS  Google Scholar 

  • El-Nahass MN, Mai EK, Ali EM (2018) Immobilization of horseradish peroxidase into cubic mesoporous silicate, SBA-16 with high activity and enhanced stability. Int J Biol Macromol 116:1304–1309

    Article  CAS  Google Scholar 

  • Esteves LC, Pinheiro AC, Pioli RM, Penna TC, Baader WJ, Correra TC, Bastos EL (2018) Revisiting the mechanism of hydrolysis of betanin. Photochem Photobiol 94:853–864

    Article  CAS  Google Scholar 

  • Gilani SL, Najafpour GD, Heydarzadeh HD, Moghadamnia A (2017) Enantioselective synthesis of (S)-naproxen using immobilized lipase on chitosan beads. Chirality 29:304–314

    Article  CAS  Google Scholar 

  • Gong A, Zhu CT, Xu Y, Wang FQ, Tsabing DK, Wu FA, Wang J (2017) Moving and unsinkable graphene sheets immobilized enzyme for microfluidic biocatalysis. Sci Rep 7:4309–4323

    Article  Google Scholar 

  • Hou CJ, Huo DQ, Mei Y, Huang S, Liang Z, Shen CH (2012) Catalytic characteristics of plant-esterase from wheat flour. World J Microbiol Biotechnol 28:541–548

    Article  CAS  Google Scholar 

  • Iyer PV, Ananthanarayan L (2008) Enzyme stability and stabilization-aqueous and non-aqueous environment. Process Biochem 43:1019–1032

    Article  CAS  Google Scholar 

  • Jiang XW, Shen TR, Tang XL, Yang WQ, Guo HH, Ling WH (2017) Cyanidin-3-O-β-glucoside combined with its metabolite protocatechuic acid attenuated the activation of mice hepatic stellate cells. Food Funct 8:2945–2957

    Article  CAS  Google Scholar 

  • Juang RS, Tseng RL, Wu FC, Lee SH (2010) Adsorption behavior of reactive dyes from aqueous solutions on chitosan. J Chem Technol Biotechnol 70:391–399

    Article  Google Scholar 

  • Khan BM, Liu ZC, Shi FL, Cheong KL, Liu Y (2018) ATPS: "aqueous two-phase system" as the "answer to protein separation" for protein-processing food industry. Crit Rev Food Sci Nutr 1–14

  • Kim DH, Sohng IS, Kobashi K, Han MJ (1996) Purification and characterization of beta-glucosidase from Bacteroides JY-6, a human intestinal bacterium. Biol Pharm Bull 19:1121–1125

    Article  CAS  Google Scholar 

  • Liu J, Peng J, Shen S, Jin QR, Li C, Yang QH (2013) Enzyme entrapped in polymer-modified nanopores: the effects of macromolecular crowding and surface hydrophobicity. Chem Eur J 19:2711–2719

    Article  CAS  Google Scholar 

  • Maryam M, Sedigheh B, Anahita H, Dorsa M, Mohammad T (2018) Immobilization of L-asparaginase on aspartic acid functionalized graphene oxide nanosheet: enzyme kinetics and stability studies. Chem Eng J 354:1153–1163

    Article  Google Scholar 

  • Natić MM, Dabić DČ, Papetti A, Fotirić MM, Ognjanov V, Ljubojević M, Tešić Ž (2015) Analysis and characterisation of phytochemicals in mulberry (Morus alba L.) fruits grown in Vojvodina, North Serbia. Food Chem 171:128–136

    Article  Google Scholar 

  • Park SY, Kim JH, Kim DH (2005) Purification and characterization of quercitrin-hydrolyzing α-L-rhamnosidase from Fusobacterium K-60, a human intestinal bacterium. J Microbiol Biotechnol 15:519–524

    CAS  Google Scholar 

  • Puskas JE, Kantor J, Shrikhande GJ (2017) Reaction engineering with enzymes: a relatively uncharted territory. AICHE J 63:266–272

    Article  CAS  Google Scholar 

  • Qin BL, Liu XC, Cui HM, Ma Y, Wang ZM, Han J (2017) Aqueous two-phase assisted by ultrasound for the extraction of anthocyanins from Lycium ruthenicum Murr. Prep Biochem Biotechnol 47:881–888

    Article  CAS  Google Scholar 

  • Rastian Z, Khodadadi AA, Vahabzadeh F (2014) Facile surface functionalization of multiwalled carbon nanotubes by soft dielectric barrier discharge plasma: generate compatible interface for lipase immobilization. Biochem Eng J 90:16–26

    Article  CAS  Google Scholar 

  • Sala L, Gautério GV, Younan FF, Brandelli A, Moraes CC, Kalil SJ (2014) Integration of ultrafiltration into an aqueous two-phase system in the keratinase purification. Process Biochem 49:2016–2024

    Article  CAS  Google Scholar 

  • Samaratunga A, Kudina O, Nahar N, Zakharchenko A, Minko S, Voronov A, Pryor SW (2015) Impact of enzyme loading on the efficacy and recovery of cellulolytic enzymes immobilized on enzymogel nanoparticles. Appl Biochem Biotechnol 175:2872–2882

    Article  CAS  Google Scholar 

  • Song HZ, Lai J, Tang Q, Zheng XD (2016) Mulberry ethanol extract attenuates hepatic steatosis and insulin resistance in high-fat diet-fed mice. Nutr Res 36:710–718

    Article  CAS  Google Scholar 

  • Suma Y, Kang CS, Kim HS (2016) Noncovalent and covalent immobilization of oxygenase on single-walled carbon nanotube for enzymatic decomposition of aromatic hydrocarbon intermediates. Environ Sci Pollut Res 23:1015–1024

    Article  CAS  Google Scholar 

  • Thomsen MS, Nidetzky B (2010) Microfluidic reactor for continuous flow biotransformations with immobilized enzymes: the example of lactose hydrolysis by a hyperthermophilic β-glycoside hydrolase. Eng Life Sci 8:40–48

    Article  Google Scholar 

  • Virgen-Ortíz JJ, Tacias-Pascacio VG, Hirata DB, Torrestiana-Sanche B, Rosales-Quintero A, Fernandez-Lafuente R (2017) Relevance of substrates and products on the desorption of lipases physically adsorbed on hydrophobic supports. Enzym Microb Technol 96:30–35

    Article  Google Scholar 

  • Wang FQ, He S, Zhu CT, Rabausch U, Streit W, Wang J (2018) The combine use of continuous-flow microchannel reactor and ionic liquid cosolvent for efficient biocatalysis of unpurified recombinant enzyme. J Chem Technol Biotechnol 93:2671–2680

    Article  CAS  Google Scholar 

  • Wang JZ, Zhu LL, Zhang F, Herman RA, Li WJ, Zhou XJ, Wu FA, Wang J (2019) Microfluidic tools for lipid production and modification: a review. Environ Sci Pollut Res 26:35482–35496

    Article  Google Scholar 

  • Wu XY, Liang LH, Zou Y, Zhao T, Zhao JL, Li F, Yang LQ (2011) Aqueous two-phase extraction, identification and antioxidant activity of anthocyanins from mulberry (Morus atropurpurea Roxb.). Food Chem 129:443–453

    Article  CAS  Google Scholar 

  • You YL, Liang C, Han X, Guo JL, Ren CL, Liu GJ, Huang WD, Zhan JC (2017) Mulberry anthocyanins, cyanidin 3-glucoside and cyanidin 3-rutinoside, increase the quantity of mitochondria during brown adipogenesis. J Funct Foods 36:348–356

    Article  CAS  Google Scholar 

  • Zhao CL, Yu YQ, Chen ZJ, Wen GS, Wei FG, Zheng Q, Wang CD, Xiao XL (2017) Stability-increasing effects of anthocyanin glycosyl acylation. Food Chem 214:119–128

    Article  CAS  Google Scholar 

  • Zhou XJ, Zhu CT, Hu Y, You S, Wu FA, Wang J (2020) A novel microfluidic aqueous two-phase system with immobilized enzyme enhances cyanidin-3-O-glucoside content in red pigments from mulberry fruits. Biochem Eng J 158:107556

    Article  CAS  Google Scholar 

Download references

Funding

This study was financially supported by Zhenjiang Science & Technology Program (Grant No. NY2017010), 333 High-level Talent Training Project of Jiangsu Province (Grant No. BRA2019281), and Shenlan Young scholars program of Jiangsu University of Science and Technology (Year 2015).

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Correspondence to Jun Wang.

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Zhou, XJ., Zhu, CT., Zhang, LY. et al. Enrichment and purification of red pigments from defective mulberry fruits using biotransformation in a liquid-liquid-solid three-phase system. Environ Sci Pollut Res 28, 24432–24440 (2021). https://doi.org/10.1007/s11356-020-08731-2

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  • DOI: https://doi.org/10.1007/s11356-020-08731-2

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