Replacing calcium with ammonium counterion in lignosulfonates from paper mills affects their molecular properties and bioactivity Davide Savy a,b, ⁎, Vincenza Cozzolino a,c , Marios Drosos a , Pierluigi Mazzei a , Alessandro Piccolo a,c a Centro Interdipartimentale di Ricerca sulla Risonanza Magnetica Nucleare per l'Ambiente, l'Agro-Alimentare ed i Nuovi Materiali (CERMANU), Università di Napoli Federico II, Via Università 100, 80055 Portici, Italy b Plant Biology laboratory, Gembloux Agro-Bio Tech, University of Liège, 2 Passage des Déportés, B-5030 Gembloux, Belgium c Dipartimento di Agraria, Università di Napoli Federico II, Via Università 100, 80055 Portici, Italy HIGHLIGHTS • Ammonium and Calcium Lignosulfo- nates (LS) were valorised as plant biostimulants. • Counterion replacement significantly affected the molecular aggregation of LS. • The materials showed different molecu- lar size and amount of sulfonates and phenols. • LS-AM positively affected the root sys- tem, LS-C increased Total and Shoot weights. • Their dose-dependent bioactivity is re- lated to their physical-chemical features. GRAPHICAL ABSTRACT abstract article info Article history: Received 21 May 2018 Received in revised form 11 July 2018 Accepted 12 July 2018 Available online xxxx Editor: Frederic Coulon Lignosulfonates are important by-products of the paper industry and may be transformed into different com- modities. We studied the molecular properties of ammonium (LS-AM) and calcium Lignosulfonates (LS-C) and evaluated their bioactivity towards the early development of maize plantlets. The FT-IR, 13 C NMR and 1 H- 13 C- HSQC-NMR spectra showed that the two lignosulfonates varied in hydroxyl, sulfonate and phenolic content, while DOSY-NMR spectroscopy suggested a similar diffusivity. High Performance Size Exclusion Chromatography (HPSEC) was used to simulate the effects of root-exuded acids and describe the conformational dynamics of both LS substrates in acidic aqueous solutions. This technique showed that LS-C was stabilized by the divalent Ca 2+ counterion, thus showing a greater conformational stability than LS-AM, whose components could not be as ef- ficiently aggregated by the monovalent NH4 + counter-ion. The plant bioassays revealed that LS-AM enhanced the elongation of the root system, whereas LS-C significantly increased both total and shoot plant weights. We concluded that the lignosulfonate bioactivity on plant growth depended on the applied concentrations, their mo- lecular properties and conformational stability. © 2018 Published by Elsevier B.V. Keywords: Plant biostimulants Water-soluble lignin NMR spectroscopy High performance size exclusion chromatogra- phy (HPSEC) Maize growth and development Intensified sustainable agriculture 1. Introduction With the progressive shortening of fossil fuels, alternative sources of fuel, energy and chemicals are searched. Lignocellulose has consider- ably attracted the interest of the scientific community (Laurichesse Science of the Total Environment 645 (2018) 411–418 ⁎ Corresponding author. E-mail address: davide.savy@unina.it (D. Savy). https://doi.org/10.1016/j.scitotenv.2018.07.153 0048-9697/© 2018 Published by Elsevier B.V. Contents lists available at ScienceDirect Science of the Total Environment journal homepage: www.elsevier.com/locate/scitotenv