, Volume 26, Issue 2, pp 1409–1415 | Cite as

Size-exclusion chromatography with on-line viscometry of various celluloses with branched and linear structures

  • Y. Ono
  • R. Funahashi
  • A. IsogaiEmail author


We analyzed softwood and hardwood bleached kraft pulps (SBKP and HBKP, respectively), and Japanese cedar (Cryptomeria japonica) celluloses prepared from wood powders using delignification, hemicellulose removal, and dilute acid hydrolysis. For sample preparation, each sample was dissolved in 8% (w/w) lithium chloride/N,N-dimethylacetamide (LiCl/DMAc), after the sample was soaked in ethylenediamine (EDA) and the EDA was exchanged with DMAc through methanol. These solutions were diluted to 1% (w/v) LiCl/DMAc and subjected to size-exclusion chromatography (SEC) combined with multiangle laser light scattering and viscometry analyses. SEC/multiangle laser light scattering and SEC/viscometry use different principles to determine the molecular structures of polymers dissolved in LiCl/DMAc. Both methods showed that SBKP celluloses with high molar masses had branched structures, whereas HBKP celluloses had linear structures as like cotton, bacterial, tunicate, and algal celluloses. Conventionally, viscosity-average molar masses or viscosity-average degrees of polymerization of SBKP and HBKP are obtained by capillary viscometry using a 0.5 M copper ethylenediamine hydroxide (cuen) solution. Because SBKP and HBKP have different cellulose structures (branched and linear molecules, respectively), their viscosity-average molar masses and viscosity-average degrees of polymerization should not be calculated using the same Mark–Houwink–Sakurada equation.

Graphical abstract


Size-exclusion chromatography with multiangle laser light scattering Size-exclusion chromatography/viscometry Mark–Houwink–Sakurada equation Softwood cellulose Cellulose molecules with branched structures 



This research was supported by the Core Research for Evolutional Science and Technology (CREST, Grant number JPMJCR13B2) of the Japan Science and Technology Agency (JST). We thank Mr. Masahide Nakamura of Shoko Co., Ltd., Tokyo, Japan, for assistance with the SEC/viscometry analyses. We thank Gabrielle David, PhD, from Edanz Group ( for editing a draft of this manuscript.


  1. Ahn S, Lee H, Lee S, Chang T (2012) Characterization of branched polymers by comprehensive two-dimensional liquid chromatography with triple detection. Macromolecules 45:3550–3556CrossRefGoogle Scholar
  2. Bikova T, Treimanis A (2002) Problems of the MMD analysis of cellulose by SEC using DMA/LiCl: a review. Carbohydr Polym 48:23–28CrossRefGoogle Scholar
  3. Berggren R, Berthold F, Sjöholm E, Lindström M (2003) Improved methods for evaluating the molar mass distributions of cellulose in kraft pulp. J Appl Polym Sci 88:1170–1179CrossRefGoogle Scholar
  4. De Gennes PG (1979) Scaling concenpts in polymer physics. Cornell University Press, Ithaca NYGoogle Scholar
  5. Dupont A, Harrison G (2004) Conformation and dn/dc determination of cellulose in N, N-dimethylacetamide containing lithium chloride. Carbohydr Polym 58:233–243CrossRefGoogle Scholar
  6. Evans SR, Wallis AFA (1987) Comparison of cellulose molecular weights determined by high performance size exclusion chromatography and viscometry. In: Proc 4th Int Symp Wood Pulping Chem, Paris, April 27–30, vol 1, pp 201–204Google Scholar
  7. Flory PJ (1953) In Principles in Polymer Chemistry. Ed by Flory PJ , Cornell University Press, Ithaca NYGoogle Scholar
  8. Hiraoki R, Ono Y, Saito T, Isogai A (2015) Molecular mass and molecular-mass distribution of TEMPO-oxidized celluloses and TEMPO-oxidized cellulose nanofibrils. Biomacromolecules 16:675–681CrossRefGoogle Scholar
  9. Kačík F, Podzimek Š, Vizárová K, Kačíková D, Čabalová I (2016) Characterization of cellulose degradation during accelerated aging by SEC-MALS, SEC-DAD, and A4F-MALS methods. Cellulose 23:357–366CrossRefGoogle Scholar
  10. Łojewski T, Zięba K, Łojewska J (2010) Size exclusion chromatography and viscometry in paper degradation studies. New Mark-Houwink coefficients for cellulose in cupri-ethylenediamine. J Chromatogr A 1217:6162–6168Google Scholar
  11. Nagy DJ (1993) A Mark-Houwink equation for polyvinyl alcohol from SEC-viscometry. J Liq Chromatogr 16:3041–3058CrossRefGoogle Scholar
  12. Oberlerchner JT, Rosenau T, Potthast A (2015) Overview of methods for the direct molar mass determination of cellulose. Molecules 20:10313–10341CrossRefGoogle Scholar
  13. Ono Y, Saito T, Isogai A (2017) Branched structures of softwood celluloses: proof based on size-exclusion chromatography and multi-angle laser-light scattering. In: Atalla RH. Agarwal UP, Isogai A (eds) Nanocelluloses: their preparation, properties, and applications, ACS Sym Ser, American Chemical Society 1251:151–169Google Scholar
  14. Ono Y, Funahashi R, Saito T, Isogai A (2018a) Investigation of stability of branched structures in softwood cellulose using SEC/MALLS/RI/UV and sugar composition analyses. Cellulose 25:2667–2679CrossRefGoogle Scholar
  15. Ono Y, Ishida T, Soeta H, Saito T, Isogai A (2016a) Reliable dn/dc values of cellulose, chitin, and cellulose triacetate dissolved in LiCl/N, N-dimethylacetamide for molecular mass analysis. Biomacromolecules 17:192–199CrossRefGoogle Scholar
  16. Ono Y, Furihata K, Isobe N, Saito T, Isogai A (2018b) Solution-state structures of the cellulose model pullulan in lithium chloride/N, N-dimethylacetamide. Int J Biol Macromol 107:2598–2603CrossRefGoogle Scholar
  17. Ono Y, Tanaka R, Funahashi R, Takeuchi M, Saito T, Isogai A (2016b) SEC–MALLS analysis of ethylenediamine-pretreated native celluloses in LiCl/N, N-dimethylacetamide: softwood kraft pulp and highly crystalline bacterial, tunicate, and algal celluloses. Cellulose 23:1639–1647CrossRefGoogle Scholar
  18. Podzimek S (2014) Truths and myths about the determination of molar mass distribution of synthetic and natural polymers by size exclusion chromatography. J Appl Polym Sci 131:40111CrossRefGoogle Scholar
  19. Podzimek S (2016) Principles of detection and characterization of branching in synthetic and natural polymers by MALS. In: Proceedings of LCGC “Beyond GPC: using light scattering for absolute polymer characterization", pp 11−16Google Scholar
  20. Podzimek S, Hermannova M, Bilerova H, Bezakova Z, Velebny V (2010) Solution properties of hyaluronic acid and comparison of SEC-MALS-VIS data with off-line capillary viscometry. J Appl Polym Sci 116:2013–3020Google Scholar
  21. Potthast A, Rosenau T, Sartori J, Sixta H, Kosma P (2003) Hydrolytic processes and condensation reactions in the cellulose solvent system N, N-dimethylacetamide/lithium chloride. Part 2: degradation of cellulose. Polymer 44:7–17CrossRefGoogle Scholar
  22. Potthast A, Radosta S, Saake B, Lebioda S, Heinze T, Henniges U, Isogai A, Koschella A, Kosma P, Rosenau T, Schiehser S, Sixta H, Strlić M, Strobin G, Vorwerg W, Wetzel H, (2015) Comparison testing of methods for gel permeation chromatography of cellulose: coming closer to a standard protocol. Cellulose 22:1591–1613CrossRefGoogle Scholar
  23. Schulz GV, Blaschke F (1941) Molecular-weight determinations on macromolecular materials. IX. An equation for the calculation of the viscosity number at very small concentrations. prakt. Chem 158:130–135Google Scholar
  24. Striegel A, Timpa JD (1995) Molecular characterization of polysaccharides dissolved in Me2NAc-LiCl by gel permeation chromatography. Carbohydr Res 267:271–290CrossRefGoogle Scholar
  25. Smith DK, Bampton RF, Alexander WJ (1963) Use of new solvents for evaluating chemical cellulose for the viscose process. Ind Eng Chem Process Design Develop 2:57–62CrossRefGoogle Scholar
  26. Shinoda R, Saito T, Okita Y, Isogai A (2012) Relationship between length and degree of polymerization of TEMPO-oxidized cellulose nanofibrils. Biomacromolecules 13:842–849CrossRefGoogle Scholar
  27. Strliča M, Kolar J, Žigon M, Pihlar B (1998) Evaluation of size-exclusion chromatography and viscometry for the determination of molecular masses of oxidised cellulose. J Chromatogr A 805:93–99CrossRefGoogle Scholar
  28. Schult T, Hjerde T, Optun OI, Kleppe PJ, Moe S (2002) Characterization of cellulose by SEC-MALLS. Cellulose 9:149–158CrossRefGoogle Scholar
  29. Test TAPPI (2009) Methods ISO/FDIS:5351Google Scholar
  30. Timpa JD (1991) Application of universal calibration in gel permeation chromatography for molecular weight determinations of plant cell wall polymers: cotton fiber. J Agrc Food Chem 39:270–275CrossRefGoogle Scholar
  31. Wise LE, Murphy M, D’Addieco AA (1946) A chlorite holocellulose, its fractionation and bearing on summative wood analysis and studies on the hemicellulose. Paper Trade J 122:35–43Google Scholar
  32. Yanagisawa M, Isogai A (2005) SEC-MALS-QELS study on the molecular conformation of celullose in LiCl/amide solutions. Biomacromolecules 6:1258–1265CrossRefGoogle Scholar
  33. Yanagisawa M, Shibata I, Isogai A (2005) SEC-MALLS analysis of softwood kraft pulp using LiCl/1,3-dimethyl-2-imidazolidinone as an eluent. Cellulose 12:151–158CrossRefGoogle Scholar
  34. Yanagisawa M, Kato Y, Yoshida Y, Isogai A (2006) SEC-MALS study on aggregates of chitosan molecules in aqueous solvents: Influence of residual N-acetyl groups. Carbohydr Polym 25:12–23Google Scholar
  35. Yamamoto M, Kuramae R, Yanagisawa M, Ishii D, Isogai A (2011) Light-scattering analysis of native wood holocelluloses totally dissolved in LiCl–DMI solutions: High probability of branched structures in inherent cellulose. Biomacromolecules 12:3982–3988CrossRefGoogle Scholar

Copyright information

© Springer Nature B.V. 2018

Authors and Affiliations

  1. 1.Department of Biomaterial SciencesThe University of TokyoTokyoJapan

Personalised recommendations