Advertisement

Journal of Fluorescence

, Volume 28, Issue 5, pp 1127–1142 | Cite as

Papain Loaded Poly(ε-Caprolactone) Nanoparticles: In-silico and In-Vitro Studies

  • Yasemin Budama-Kilinc
  • Rabia Cakir-Koc
  • Serda Kecel-Gunduz
  • Tolga Zorlu
  • Yagmur Kokcu
  • Bilge Bicak
  • Zeynep Karavelioglu
  • Aysen E. Ozel
ORIGINAL ARTICLE
  • 159 Downloads

Abstract

Papain is a protease enzyme with therapeutic properties that are very valuable for medical applications. Poly(ε-caprolactone) (PCL) is an ideal polymeric carrier for controlled drug delivery systems due to its low biodegradability and its high biocompatibility. In this study, the three-dimensional structure and action mechanism of papain were investigated by in vitro and in silico experiments using molecular dynamics (MD) and molecular docking methods to elucidate biological functions. The results showed that the size of papain-loaded PCL nanoparticles (NPs) and the polydispersity index (PDI) of the NPs were 242.9 nm and 0.074, respectively. The encapsulation efficiency and loading efficiency were 80.4 and 27.2%, respectively. Human embryonic kidney cells (HEK-293) were used for determining the cytotoxicity of papain-loaded PCL and PCL nanoparticles. The in vitro cell culture showed that nanoparticles are not toxic at low concentrations, while toxicity slightly increases at high concentrations. In silico studies, which were carried out with MD simulations and ADME analysis showed that the strong hydrogen bonds between the ligand and the papain provide stability and indicate the regions in which the interactions occur.

Keywords

Papain Enzyme Fluorescence Imaging PCL HEK-293 Molecular docking Molecular dynamic 

Notes

Acknowledgments

Authors are very thankful Rita Podzuna for allowing to use the docking program with Schrödinger's Small-Molecule Drug Discory Suite.

Compliance with Ethical Standards

Conflict of Interest

The authors declare that there are no conflicts of interest associated with this work.

Supplementary material

10895_2018_2276_Fig19_ESM.png (470 kb)
ESM 1

(PNG 469 kb)

10895_2018_2276_MOESM1_ESM.tif (1 mb)
Hıgh Resolutıon Image (TIF 1073 kb)
10895_2018_2276_Fig20_ESM.png (421 kb)
ESM 2

(PNG 420 kb)

10895_2018_2276_MOESM2_ESM.tif (1.2 mb)
Hıgh Resolutıon Image (TIF 1225 kb)
10895_2018_2276_Fig21_ESM.png (511 kb)
ESM 3

(PNG 510 kb)

10895_2018_2276_MOESM3_ESM.tif (1.7 mb)
Hıgh Resolutıon Image (TIF 1704 kb)
10895_2018_2276_Fig22_ESM.png (571 kb)
ESM 4

(PNG 571 kb)

10895_2018_2276_MOESM4_ESM.tif (1.7 mb)
Hıgh Resolutıon Image (TIF 1757 kb)
10895_2018_2276_Fig23_ESM.png (758 kb)
ESM 5

(PNG 757 kb)

10895_2018_2276_MOESM5_ESM.tif (1.8 mb)
Hıgh Resolutıon Image (TIF 1885 kb)
10895_2018_2276_Fig24_ESM.png (340 kb)
ESM 6

(PNG 339 kb)

10895_2018_2276_MOESM6_ESM.tif (902 kb)
Hıgh Resolutıon Image (TIF 902 kb)

References

  1. 1.
    Liang Y-Y, Zhang L-M (2007) Bioconjugation of papain on superparamagnetic nanoparticles decorated with carboxymethylated chitosan. Biomacromolecules 8(5):1480–1486CrossRefPubMedGoogle Scholar
  2. 2.
    Peng J, Han CL, Ling J, Liu CJ, Ding ZT, Cao QE (2018) Selective fluorescence quenching of papain–au nanoclusters by self-polymerization of dopamine. Luminescence 33(1):168–173CrossRefPubMedGoogle Scholar
  3. 3.
    Mamboya EAF (2012) Papain, a plant enzyme of biological importance: a review. Am J Biochem Biotechnol 8(2):99–104CrossRefGoogle Scholar
  4. 4.
    dos Anjos MM, da Silva AA, de Pascoli IC, Mikcha JMG, Machinski M Jr, Peralta RM, de Abreu Filho BA (2016) Antibacterial activity of papain and bromelain on Alicyclobacillus spp. Int J Food Microbiol 216:121–126CrossRefPubMedGoogle Scholar
  5. 5.
    Silva CRd, Oliveira MB, Motta ES, Almeida GSd, Varanda LL, Pádula Md, Leitão AC, Caldeira-de-Araújo A (2010) Genotoxic and cytotoxic safety evaluation of papain (Carica papaya L.) using in vitro assays. Biomed Res Int 2010Google Scholar
  6. 6.
    Xiao Q, Qiu H, Huang S, Huang C, Su W, Hu B, Liu Y (2013) Systematic investigation of interactions between papain and MPA-capped CdTe quantum dots. Mol Biol Rep 40(10):5781–5789CrossRefPubMedGoogle Scholar
  7. 7.
    Adebiyi A, Adaikan PG, Prasad R (2002) Papaya (Carica papaya) consumption is unsafe in pregnancy: fact or fable? Scientific evaluation of a common belief in some parts of Asia using a rat model. Br J Nutr 88(2):199–203CrossRefPubMedGoogle Scholar
  8. 8.
    Mansfield LE, Bowers CH (1983) Systemic reaction to papain in a nonoccupational setting. J Allergy Clin Immunol 71(4):371–374CrossRefPubMedGoogle Scholar
  9. 9.
    Vasconcellos FC, Goulart GA, Beppu MM (2011) Production and characterization of chitosan microparticles containing papain for controlled release applications. Powder Technol 205(1–3):65–70CrossRefGoogle Scholar
  10. 10.
    Chen Y-Y, Lu Y-H, Ma C-H, Tao W-W, Zhu J-J, Zhang X (2017) A novel elastic liposome for skin delivery of papain and its application on hypertrophic scar. Biomed Pharmacother 87:82–91CrossRefPubMedGoogle Scholar
  11. 11.
    Alkilani AZ, McCrudden MT, Donnelly RF (2015) Transdermal drug delivery: innovative pharmaceutical developments based on disruption of the barrier properties of the stratum corneum. Pharmaceutics 7(4):438–470CrossRefPubMedGoogle Scholar
  12. 12.
    Herman A, Herman AP (2015) Essential oils and their constituents as skin penetration enhancer for transdermal drug delivery: a review. J Pharm Pharmacol 67(4):473–485CrossRefPubMedGoogle Scholar
  13. 13.
    Mout R, Ray M, Tay T, Sasaki K, Yesilbag Tonga G, Rotello VM (2017) General strategy for direct cytosolic protein delivery via protein–nanoparticle co-engineering. ACS Nano 11(6):6416–6421CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Maximov V, Reukov V, Vertegel A (2009) Targeted delivery of therapeutic enzymes. J Drug Delivery Sci Technol 19(5):311–320CrossRefGoogle Scholar
  15. 15.
    Woodruff MA, Hutmacher DW (2010) The return of a forgotten polymer—polycaprolactone in the 21st century. Prog Polym Sci 35(10):1217–1256CrossRefGoogle Scholar
  16. 16.
    Smith PE, van Gunsteren WF (1993) The viscosity of SPC and SPC/E water at 277 and 300 K. Chem Phys Lett 215(4):315–318CrossRefGoogle Scholar
  17. 17.
    Fletcher R (2001) Practical methods of optimization, 2nd edn. Wiley, Chichester, 2000. Numerical Algorithms 26 (2):198Google Scholar
  18. 18.
    Parrinello M, Rahman A (1981) Polymorphic transitions in single crystals: a new molecular dynamics method. J Appl Phys 52(12):7182–7190CrossRefGoogle Scholar
  19. 19.
    Berendsen HJ, Jv P, van Gunsteren WF, DiNola A, Haak J (1984) Molecular dynamics with coupling to an external bath. J Chem Phys 81(8):3684–3690CrossRefGoogle Scholar
  20. 20.
    Hess B, Bekker H, Berendsen HJ, Fraaije JG (1997) LINCS: a linear constraint solver for molecular simulations. J Comput Chem 18(12):1463–1472CrossRefGoogle Scholar
  21. 21.
    Darden T, York D, Pedersen L (1993) Particle mesh Ewald: an N· log (N) method for Ewald sums in large systems. J Chem Phys 98(12):10089–10092CrossRefGoogle Scholar
  22. 22.
    Verlet L (1967) Computer" experiments" on classical fluids. I. Thermodynamical properties of Lennard-Jones molecules. Phys Rev 159(1):98CrossRefGoogle Scholar
  23. 23.
    Van Der Spoel D, Lindahl E, Hess B, Groenhof G, Mark AE, Berendsen HJ (2005) GROMACS: fast, flexible, and free. J Comput Chem 26(16):1701–1718CrossRefGoogle Scholar
  24. 24.
    van Gunsteren WF, Billeter SR, Eising AA, Hünenberger PH, Krüger P, Mark AE, Scott WR, Tironi IG (1996) Biomolecular simulation: the {GROMOS96} manual and user guideGoogle Scholar
  25. 25.
    Humphrey W, Dalke A, Schulten K (1996) VMD: visual molecular dynamics. J Mol Graph 14(1):33–38CrossRefPubMedGoogle Scholar
  26. 26.
    Friesner RA, Murphy RB, Repasky MP, Frye LL, Greenwood JR, Halgren TA, Sanschagrin PC, Mainz DT (2006) Extra precision glide: docking and scoring incorporating a model of hydrophobic enclosure for protein− ligand complexes. J Med Chem 49(21):6177–6196CrossRefPubMedGoogle Scholar
  27. 27.
    Halgren TA, Murphy RB, Friesner RA, Beard HS, Frye LL, Pollard WT, Banks JL (2004) Glide: a new approach for rapid, accurate docking and scoring. 2. Enrichment factors in database screening. J Med Chem 47(7):1750–1759CrossRefPubMedGoogle Scholar
  28. 28.
    Friesner RA, Banks JL, Murphy RB, Halgren TA, Klicic JJ, Mainz DT, Repasky MP, Knoll EH, Shelley M, Perry JK (2004) Glide: a new approach for rapid, accurate docking and scoring. 1. Method and assessment of docking accuracy. J Med Chem 47(7):1739–1749CrossRefPubMedGoogle Scholar
  29. 29.
    Tsuge H, Nishimura T, Tada Y, Asao T, Turk D, Turk V, Katunuma N (1999) Inhibition mechanism of cathepsin L-specific inhibitors based on the crystal structure of papain–CLIK148 complex. Biochem Biophys Res Commun 266(2):411–416CrossRefPubMedGoogle Scholar
  30. 30.
    Bienert S, Waterhouse A, de Beer TA, Tauriello G, Studer G, Bordoli L, Schwede T (2016) The SWISS-MODEL repository—new features and functionality. Nucleic Acids Res 45(D1):D313–D319CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    Søndergaard CR, Olsson MH, Rostkowski M, Jensen JH (2011) Improved treatment of ligands and coupling effects in empirical calculation and rationalization of p K a values. J Chem Theory Comput 7(7):2284–2295CrossRefPubMedGoogle Scholar
  32. 32.
    Sastry GM, Adzhigirey M, Day T, Annabhimoju R, Sherman W (2013) Protein and ligand preparation: parameters, protocols, and influence on virtual screening enrichments. J Comput Aided Mol Des 27(3):221–234CrossRefPubMedGoogle Scholar
  33. 33.
    Harder E, Damm W, Maple J, Wu C, Reboul M, Xiang JY, Wang L, Lupyan D, Dahlgren MK, Knight JL (2015) OPLS3: a force field providing broad coverage of drug-like small molecules and proteins. J Chem Theory Comput 12(1):281–296CrossRefPubMedGoogle Scholar
  34. 34.
    Hirt RP, de Miguel N, Nakjang S, Dessi D, Liu Y-C, Diaz N, Rappelli P, Acosta-Serrano A, Fiori P-L, Mottram JC (2011) Trichomonas vaginalis pathobiology: new insights from the genome sequence. In: advances in parasitology, vol 77. Elsevier, pp 87-140Google Scholar
  35. 35.
    Carlton JM, Hirt RP, Silva JC, Delcher AL, Schatz M, Zhao Q, Wortman JR, Bidwell SL, Alsmark UCM, Besteiro S (2007) Draft genome sequence of the sexually transmitted pathogen trichomonas vaginalis. Science 315(5809):207–212CrossRefPubMedPubMedCentralGoogle Scholar
  36. 36.
    Setzer MS, Byler KG, Ogungbe IV, Setzer WN (2017) Natural products as new treatment options for trichomoniasis: a molecular docking investigation. Sci Pharm 85(1):5CrossRefPubMedCentralGoogle Scholar
  37. 37.
    Crane SN, Black WC, Palmer JT, Davis DE, Setti E, Robichaud J, Paquet J, Oballa RM, Bayly CI, McKay DJ (2006) β-Substituted cyclohexanecarboxamide: a nonpeptidic framework for the design of potent inhibitors of cathepsin K. J Med Chem 49(3):1066–1079CrossRefPubMedGoogle Scholar
  38. 38.
    Epple R, Urbina HD, Russo R, Liu H, Mason D, Bursulaya B, Tumanut C, Li J, Harris JL (2007) Bicyclic carbamates as inhibitors of papain-like cathepsin proteases. Bioorg Med Chem Lett 17(5):1254–1259CrossRefPubMedGoogle Scholar
  39. 39.
    Lipinski C, Lombardo F, Dominy B, Feeney P (1997) Toward minimalistic modeling of oral drug absorption. Adv Drug Deliv Rev 23:3–25CrossRefGoogle Scholar
  40. 40.
    Elzein T, Nasser-Eddine M, Delaite C, Bistac S, Dumas P (2004) FTIR study of polycaprolactone chain organization at interfaces. J Colloid Interface Sci 273(2):381–387CrossRefPubMedGoogle Scholar
  41. 41.
    Abdelrazek EM, Hezma AM, El-khodary A, Elzayat AM (2016) Spectroscopic studies and thermal properties of PCL/PMMA biopolymer blend. Egypt J Basic Appl Sci 3(1):10–15CrossRefGoogle Scholar
  42. 42.
    Liu K, Kiran E (2008) High-pressure solution blending of poly (ε-caprolactone) with poly (methyl methacrylate) in acetone + carbon dioxide. Polymer 49:1555–1561CrossRefGoogle Scholar
  43. 43.
    Tang ZG, Black RA, Curran JM, Hunt JA, Rhodes NP, Williams DF (2004) Surface properties and biocompatibility of solvent-cast poly [ε-caprolactone] films. Biomaterials 25(19):4741–4748CrossRefPubMedGoogle Scholar
  44. 44.
    Catledge SA, Clem WC, Shrikishen N, Chowdhury S, Stanishevsky AV, Koopman M, Vohra YK (2007) An electrospun triphasic nanofibrous scaffold for bone tissue engineering. Biomed Mater 2(2):142–150CrossRefPubMedGoogle Scholar
  45. 45.
    Goormaghtigh E, Cabiaux V, Ruysschaert JM (1990) Secondary structure and dosage of soluble and membrane proteins by attenuated total reflection Fourier‐transform infrared spectroscopy on hydrated films. FEBS J 193(2):409–420Google Scholar
  46. 46.
    Vedantham G, Sparks HG, Sane SU, Tzannis S, Przybycien TM (2000) A holistic approach for protein secondary structure estimation from infrared spectra in H2O solutions. Anal Biochem 285(1):33–49CrossRefPubMedGoogle Scholar
  47. 47.
    Goossens K, Haelewyn J, Meersman F, De Ley M, Heremans K (2003) Pressure-and temperatureinduced unfolding and aggregation of recombinant human interferon-gamma: a Fourier transform infrared spectroscopy study. Biochem J 370(2):529–535CrossRefPubMedPubMedCentralGoogle Scholar
  48. 48.
    Ismail AA, Mantsch HH, Wong PT (1992) Aggregation of chymotrypsinogen: portrait by infrared spectroscopy. Biochim Biophys Acta Protein Struct Mol Enzymol 1121(1-2):183–188CrossRefGoogle Scholar
  49. 49.
    Sharma M, Sharma V, Panda AK, Majumdar DK (2013) Development of enteric submicron particles formulation of α-amylase for oral delivery. Pharm Dev Technol 18(3):560–569CrossRefPubMedGoogle Scholar
  50. 50.
    Forato LA, Bernardes-Filho R, Colnago LA (1998) Protein structure in KBr pellets by infrared spectroscopy. Anal Biochem 259(1):136–141CrossRefPubMedGoogle Scholar
  51. 51.
    Mahmoud KA, Lam E, Hrapovic S, Luong JH (2013) Preparation of well-dispersed gold/magnetite nanoparticles embedded on cellulose nanocrystals for efficient immobilization of papain enzyme. ACS Appl Mater Interfaces 5(11):4978–4985CrossRefPubMedGoogle Scholar
  52. 52.
    Smith BC (1998) Infrared spectral interpretation: a systematic approach. CRC pressGoogle Scholar
  53. 53.
    Heimowska A, Morawska M, Bocho-Janiszewska A (2017) Biodegradation of poly (ε-caprolactone) in natural water environments. Pol J Chem Technol 19(1):120–126CrossRefGoogle Scholar
  54. 54.
    López-García J, Lehocký M, Humpolíček P, Sáha P (2014) HaCaT keratinocytes response on antimicrobial atelocollagen substrates: extent of cytotoxicity, cell viability and proliferation. J Funct Biomater 5(2):43–57CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Yasemin Budama-Kilinc
    • 1
  • Rabia Cakir-Koc
    • 1
  • Serda Kecel-Gunduz
    • 2
  • Tolga Zorlu
    • 3
  • Yagmur Kokcu
    • 4
  • Bilge Bicak
    • 2
    • 4
  • Zeynep Karavelioglu
    • 3
  • Aysen E. Ozel
    • 2
  1. 1.Faculty of Chemical and Metallurgical Engineering, Department of BioengineeringYildiz Technical UniversityIstanbulTurkey
  2. 2.Faculty of Science, Physics DepartmentIstanbul UniversityIstanbulTurkey
  3. 3.Graduate School of Natural and Applied ScienceYildiz Technical UniversityIstanbulTurkey
  4. 4.Graduate School of Engineering and SciencesIstanbul UniversityIstanbulTurkey

Personalised recommendations