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Single-step purification of C-phycocyanin from Arthrospira platensis using aqueous two-phase system based on natural deep eutectic solvents

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Abstract

Natural deep eutectic solvents (NADESs) are an emerging type of green ionic liquid that has been used in aqueous two-phase systems (ATPSs). C-phycocyanin (C-PC) is an important fluorescent protein found in cyanobacteria with multiple applications. In this study, NADES [choline chloride-urea, ChCl-U]-ATPS (NADES-ATPS) was screened as an optimum system for extracting C-PC from Arthrospira platensis. Under the optimal purification conditions (crude extracted C-PC, 500 μg; [ChCl-U], 3.25 g; K2HPO4, 0.90 g mL−1; extraction time, 2 h; NaCl, 2% and 25 °C), C-PC with a purity and recovery of 3.383 and 65.64%, respectively, was obtained by single-step extraction. The purity of C-PC could increase to 4.100 after four extraction steps. The purified C-PC was further analyzed via sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), which showed α and β subunits with molecular weights of 18.0 and 21.0 kDa, respectively. Importantly, the NADES can successfully be recycled and reused for at least five pretreatment cycles. Furthermore, no structural changes in C-PC were detected, and the aggregation phenomenon of NADES with C-PC may be the extraction mechanism. Such a green, easily synthesized, recyclable and efficient method using NADES-ATPS could hold promise for wide applications for the purification of C-PC and other proteins.

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References

  • Abd El Baky HH, El Baroty GS, Ibrahem EA (2015) Functional characters evaluation of biscuits sublimated with pure phycocyanin isolated from Spirulina and Spirulina biomass. Nutr Hosp 32:231–241

    CAS  PubMed  Google Scholar 

  • Andel F, Murphy JT, Haas JA, McDowell MT, van der Hoef I, Lugtenburg J, Lagarias JC, Mathies RA (2000) Probing the photoreaction mechanism of phytochrome through analysis of resonance Raman vibrational spectra of recombinant analogues. Biochemistry 39:2667–2676

    CAS  PubMed  Google Scholar 

  • Bai CJ, Wei QF, Ren XL (2017) Selective extraction of collagen peptides with high purity from cod skins by deep eutectic solvents. ACS Sustain Chem Eng 5:7220–7227

    CAS  Google Scholar 

  • Benavides J, Rito-Palomares M (2005) Potential aqueous two-phase processes for the primary recovery of colored protein from microbial origin. Eng Life Sci 5:259–266

    CAS  Google Scholar 

  • Chen Z, Reznicek WD, Wan CX (2018) Deep eutectic solvent pretreatment enabling full utilization of switchgrass. Bioresour Technol 263:40–48

    CAS  PubMed  Google Scholar 

  • Chethana S, Nayak CA, Madhusudhan MC, Raghavarao KSMS (2015) Single step aqueous two-phase extraction for downstream processing of C-phycocyanin from Spirulina platensis. J Food Sci Technol 52:2415–2421

    CAS  PubMed  Google Scholar 

  • Cian RE, López-Posadas R, Drago SR, de Medina FS, Martínez-Augustin O (2012) Immunomodulatory properties of the protein fraction from Phorphyra columbina. J Agric Food Chem 60:8146–8154

    CAS  PubMed  Google Scholar 

  • Coates J (2006) Interpretation of infrared spectra, a practical approach. John Wiley and Sons Ltd., Chichester

    Google Scholar 

  • Dai YT, Witkamp GJ, Verpoorte R, Choi YH (2013) Natural deep eutectic solvents as a new extraction media for phenolic metabolites in Carthamus tinctorius L. Anal Chem 85:6272–6278

    CAS  PubMed  Google Scholar 

  • Devi AC, Tavanandi HA, Govindaraju K, Raghavarao KSMS (2020) An effective method for extraction of high purity phycocyanins (C-PC and A-PC) from dry biomass of Arthrospira maxima. J Appl Phycol 32:1141–1151

    CAS  Google Scholar 

  • Figueira FD, Moraes CC, Kalil SJ (2018) C-phycocyanin purification: multiple processes for different applications. Braz J Chem Eng 35:1117–1128

    CAS  Google Scholar 

  • Hou YH, Yan MH, Wang QF, Wang YF, Xu YF, Wang YT, Li HY, Wang H (2017) C-phycocyanin from Spirulina maxima as a green fluorescent probe for the highly selective detection of mercury(II) in seafood. Food Anal Methods 10:1931–1939

    Google Scholar 

  • Jiang Z, Yuan JG, Fan XR, Xu J, Wang K, Zhang LB (2018) Dissolution and regeneration of wool keratin in the deep eutectic solvent of choline chloride-urea. Int J Biol Macromol 119:423–430

    CAS  PubMed  Google Scholar 

  • Kneip C, Hildebtandt P, Németh K, Mark F, Schaffner K (1999) Interpretation of the resonance Raman spectra of linear tetrapyrroles based on DFT calculations. Chem Phys Lett 31:479–484

    Google Scholar 

  • Kovač D, Babić O, Milovanović I, Mišan A, Simeunović J (2017) The production of biomass and phycobiliprotein pigments in filamentous cyanobacteria: the impact of light and carbon sources. Appl Biochem Biotechnol 53:539–545

    Google Scholar 

  • Kumar D, Dhar DW, Pabbi S, Kumar N, Walia S (2014) Extraction and purification of C-phycocyanin from Spirulina platensis (CCC540). J Plant Physiol 2014:184–188

    Google Scholar 

  • Kumar AK, Parikh BS, Pravakar M (2016) Natural deep eutectic solvent mediated pretreatment of rice straw: bioanalytical characterization of lignin extract and enzymatic hydrolysis of pretreated biomass residue. Environ Sci Pollut Res 23:9265–9275

    CAS  Google Scholar 

  • Li N, Wang YZ, Xu KJ, Huang YH, Wen Q, Ding XQ (2016a) Development of green betaine-based deep eutectic solvent aqueous two-phase system for the extraction of protein. Talanta 152:23–32

    CAS  PubMed  Google Scholar 

  • Li N, Wang YZ, Xu JK, Wen Q, Ding XQ, Zhang HM, Yang Q (2016b) High-performance of deep eutectic solvent based aqueous bi-phasic systems for the extraction of DNA. RSC Adv 6:84406–84414

    CAS  Google Scholar 

  • Liao XX, Zhang BC, Wang XQ, Yan HD, Zhang XW (2011) Purification of C-phycocyanin from Spirulina platensis by single-step ion-exchange chromatography. Chromatographia 73:91–296

    Google Scholar 

  • Liu Y, Feng YQ, Lun JM (2012) Aqueous two-phase countercurrent distribution for the separation of C-phycocyanin and allophycocyanin from Spirulina platensis. Food Bioprod Process 90:111–117

    CAS  Google Scholar 

  • Luo X, Smith P, Raston CL, Zhang W (2016) Vortex fluidic device-intensified aqueous two phase extraction of C-phycocyanin from Spirulina maxima. ACS Sustain Chem Eng 4:3905–3911

    CAS  Google Scholar 

  • Meng JJ, Wang YZ, Zhou YG, Chen J, Wei XX, Ni R, Liu ZW, Xu FT (2019) Development of different deep eutectic solvent aqueous biphasic systems for the separation of proteins. RSC Adv 9:14116–14125

    CAS  Google Scholar 

  • Mogany T, Kumari S, Swalaha FM, Bux F (2019) Extraction and characterisation of analytical grade C-phycocyanin from Euhalothece sp. J Appl Phycol 31:1661–1674

    CAS  Google Scholar 

  • Nganou C, David L, Meinke R, Adir N, Maultzsch J, Mkandawire M, Pouhè D, Thomsen C (2014) Activation and deactivation of vibronic channels in intact phycocyanin rods. J Chem Phys 140:085101

    CAS  PubMed  Google Scholar 

  • Pal CBT, Jadeja GC (2019) Deep eutectic solvent-based extraction of polyphenolic antioxidants from onion (Allium cepa L.) peel. J Sci Food Agric 99:1969–1979

    CAS  PubMed  Google Scholar 

  • Pandey A, Pandey S (2014) Solvatochromic probe behavior within choline chloride-based deep eutectic solvents: effect of temperature and water. J Phys Chem B 118:14652–14661

    CAS  PubMed  Google Scholar 

  • Pang JY, Sha XF, Chao YH, Chen GY, Han CR, Zhu WS, Li HM, Zhang Q (2017) Green aqueous biphasic systems containing deep eutectic solvents and sodium salts for the extraction of protein. RSC Adv 7:49361–49367

    CAS  Google Scholar 

  • Park JH, Oh KW, Choi HM (2013) Preparation and characterization of cotton fabrics with antibacterial properties treated by crosslinkable benzophenone derivative in choline chloride-based deep eutectic solvents. Cellulose 20:2101–2114

    CAS  Google Scholar 

  • Patel HM, Rastogi RP, Trivedi U, Madamwar D (2018) Structural characterization and antioxidant potential of phycocyanin from the cyanobacterium Geitlerinema sp. H8DM. Algal Res 32:372–383

    Google Scholar 

  • Patil G, Raghavarao KSMS (2007) Aqueous two phase extraction for purification of C-phycocyanin. Biochem Eng J 34:156–164

    CAS  Google Scholar 

  • Piemontese L, Perna FM, Logrieco A, Capriati V, Solfrizzo M (2017) Deep eutectic solvents as novel and effective extraction media for quantitative determination of ochratoxin A in wheat and derived products. Molecules 22:121

    PubMed Central  Google Scholar 

  • Rodríguez-Sánchez R, Ortiz-Butrón R, Blas-Valdivia V, Hernández-García A, Cano-Europa E (2012) Phycobiliproteins or C-phycocyanin of Arthrospira (Spirulina) maxima protect against HgCl2-caused oxidative stress and renal damage. Food Chem 135:2359–2365

    PubMed  Google Scholar 

  • Saini MK, Sanyal SN (2015) Cell cycle regulation and apoptotic cell death in experimental colon carcinogenesis: intervening with cyclooxygenase-2 inhibitors. Nutr Cancer 67:620–636

    CAS  PubMed  Google Scholar 

  • Silva LA, Kuhn KR, Moraes CC, Burkert CAV, Kalil SJ (2009) Experimental design as a tool for optimization of C-phycocyanin purification by precipitation from Spirulina platensis. J Braz Chem Soc 20:5–12

    CAS  Google Scholar 

  • Silveira ST, Quines LKDM, Burkert CAV, Kalil SJ (2008) Separation of phycocyanin from Spirulina platensis using ion exchange chromatography. Bioprocess Biosyst Eng 31:477–482

    CAS  PubMed  Google Scholar 

  • Singh NK, Sonani RR, Rastogi RP, Madamwar D (2015) The phycobilisomes: an early requisite for efficient photosynthesis in cyanobacteria. EXCLI J 14:268–289

    PubMed  PubMed Central  Google Scholar 

  • Steichen M, Thomassey M, Siebentritt S, Dale PJ (2011) Controlled electrodeposition of Cu-Ga from a deep eutectic solvent for low cost fabrication of CuGaSe2 thin film solar cells. Phys Chem Chem Phys 13:4292–4302

    CAS  PubMed  Google Scholar 

  • Surewicz WK, Mantsch HH, Chapman D (1993) Determination of protein secondary structure by Fourier transform infrared spectroscopy: a critical assessment. Biochemistry 32:389–394

    CAS  PubMed  Google Scholar 

  • Thangam R, Suresh V, Princy WA, Rajkumar M, SenthilKumar N, Gunasekaran P, Rengasamy R, Anbazhagan C, Kaveri K, Kannan S (2013) C-phycocyanin from Oscillatoria tenuis exhibited an antioxidant and in vitro antiproliferative activity through induction of apoptosis and G0/G1 cell cycle arrest. Food Chem 140:262–272

    CAS  PubMed  Google Scholar 

  • Wang S, Peng XW, Zhong LX, Jing SS, Cao XF, Lu FC, Sun RC (2015) Choline chloride/urea as an effective plasticizer for production of cellulose films. Carbohydr Polym 117:133–139

    CAS  PubMed  Google Scholar 

  • Wang T, Xu WJ, Wang SX, Kou P, Wang P, Wang XQ, Fu YJ (2017) Integrated and sustainable separation of chlorogenic acid from blueberry leaves by deep eutectic solvents coupled with aqueous two-phase system. Food Bioprod Process 107:205–214

    Google Scholar 

  • Wang LT, Yang Q, Cui Q, Fan XH, Dong MZ, Gao MZ, Lv MJ, An JY, Meng D, Zhao XH, Fu YJ (2020) Recyclable menthol-based deep eutectic solvent micellar system for extracting phytochemicals from Ginkgo biloba leaves. J Clean Prod 244:118648

    CAS  Google Scholar 

  • Xu KJ, Wang YZ, Huang YH, Li N, Wen Q (2015) A green deep eutectic solvent-based aqueous two-phase system for protein extracting. Anal Chim Acta 864:9–20

    CAS  PubMed  Google Scholar 

  • Xu PL, Wang YZ, Chen J, Wei XX, Xu W, Ni R, Meng JJ, Zhou YG (2018) A novel aqueous biphasic system formed by deep eutectic solvent and ionic liquid for DNA partitioning. Talanta 189:467–479

    CAS  PubMed  Google Scholar 

  • Xu PL, Wang YZ, Chen J, Wei XX, Xu W, Ni R, Meng JJ (2019) Development of deep eutectic solvent-based aqueous biphasic system for the extraction of lysozyme. Talanta 202:1–10

    CAS  PubMed  Google Scholar 

  • Xu KJ, Xu PL, Wang YZ (2020) Aqueous biphasic systems formed by hydrophilic and hydrophobic deep eutectic solvents for the partitioning of dyes. Talanta 213:120839

    CAS  PubMed  Google Scholar 

  • Zeng Q, Wang YZ, Huang YH, Ding XQ, Chen J, Xu KJ (2014) Deep eutectic solvents as novel extraction media for protein partitioning. Analyst 139:2565–2573

    CAS  PubMed  Google Scholar 

  • Zhang HM, Wang YZ, Xu KJ, Li N, Wen Q, Yang Q, Zhou YG (2016) Ternary and binary deep eutectic solvents as a novel extraction media for protein partitioning. Anal Methods-UK 8:8196–8207

    CAS  Google Scholar 

  • Zhao L, Peng YL, Gao JM, Cai WM (2014) Bioprocess intensification: an aqueous two-phase process for the purification of C-phycocyanin from dry Spirulina platensis. Eur Food Res Technol 238:451–457

    CAS  Google Scholar 

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Funding

This work was supported by the National Natural Science Foundation of China, China (41876149); the Key Research and Development Plan of Shandong Province, China (2019GHY112031); and the Natural Science Foundation of Shandong Province, China (ZR2019MD018).

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Authors and Affiliations

  1. School of Marine Science and Technology, Harbin Institute of Technology, Weihai, 264209, People’s Republic of China

    Quanfu Wang, Nana Wei, Yifan Wang, Yanhua Hou, Xiulian Ren & Qifeng Wei

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  1. Quanfu Wang
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  2. Nana Wei
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  3. Yifan Wang
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  4. Yanhua Hou
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  5. Xiulian Ren
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Correspondence to Yanhua Hou.

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Wang, Q., Wei, N., Wang, Y. et al. Single-step purification of C-phycocyanin from Arthrospira platensis using aqueous two-phase system based on natural deep eutectic solvents. J Appl Phycol 32, 3873–3883 (2020). https://doi.org/10.1007/s10811-020-02239-5

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