Lignin derivatives stabilizing oil-in-water emulsions: Technological aspects, interfacial rheology and cytotoxicity

https://doi.org/10.1016/j.indcrop.2020.112762Get rights and content

Highlights

  • Calcium lignosulfonate and kraft lignin were more efficient in decreasing the interfacial tension than whey protein isolate.

  • Lignins formed a denser and more elastic interfacial layer than whey protein isolate (WPI).

  • Emulsions produced with lignins were more stable than those produced with WPI during 28 days.

  • Stable emulsions were produced with calcium lignosulfonate and kraft lignin at the safety level of cytotoxicity.

Abstract

The emulsifier ability to stabilize oil-in-water emulsions of two industrial lignins (calcium lignosulfonate and kraft lignin) were evaluated and compared with whey protein isolate that is a widely used emulsifier, as well as their cytotoxic effect in Caco-2 and HT-29 cell lines. The surface activity of calcium lignosulfonate (LG) and kraft lignin (LA) was evidenced by the reduction of interfacial tension between oil-water phases, and interfacial activity was lower and presented a faster decrease than those obtained for whey protein. This behavior was associated to the great driving force promoted by the high number of hydrophobic groups on the lignins. The emulsions with lignins presented smaller diameter (0.717 μm) and higher zeta potential than emulsion produced with WPI (1.065 μm). In addition, WPI emulsions were the most unstable after 28 days of storage. Among lignins, emulsions with kraft lignin were the most stable due to differences in the functional groups on these lignins. The cytotoxicity of lignins towards HT-29 and Caco-2 cells showed a dose-response relationship, determining non-cytotoxic concentrations for emulsifiers that can produce stable emulsions. In order to encourage the industries to use these lignins in polymeric form, these results present the safety level for both lignins capable of producing a stable emulsion, in addition to presenting characteristics of these two biopolymers that will influence the different applications.

Introduction

Emulsions have been widely used to protect, vehiculate, and deliver lipophilic bioactive molecules, since the bioaccessibility of these functional compounds can increase if they are carried in O/W emulsions (McClements, 2015). One of the critical points in the production of a stable emulsion is the choice of a suitable emulsifier, but due to the increase in consumer demand for environmental preservation and health, the industry is looking for new natural emulsifiers. Although several biodegradable ingredients (proteins, polysaccharides, and phospholipids) are used to stabilize emulsions, synthetic emulsifiers are still widely employed because of their high emulsifying efficiency (Ozturk and McClements, 2016; Riquelme et al., 2019).

A material is considered adequate for industrial application as an additive if it is available in large quantity, presents low cost and good technological properties, but also shows low toxicity (Shulga et al., 2011). Lignin is an abundant plant-derived material and a waste of the pulp, paper, ethanol, textile, and food industry. Approximately 100 million tons of lignin are produced annually, but less than 2% is used in commercial products with higher added value. The rest of the production is used as fuel for boilers or is discarded (Aro and Fatehi, 2017; Bajwa et al., 2019; Lievonen et al., 2016). Lignin is the second most common biopolymer of vegetable origin, and it is the unique renewable source material that has an aromatic structure. Additionally, this polymer is biodegradable and present high thermal stability, antioxidant, antimicrobial, antifungal, and anti-cancer properties. Therefore, identification of technological properties of lignins as emulsifier is relevant to add value to this waste biopolymer for cleaning, food, personal care, cosmetic, paint, biomedical, chemical, agricultural and pharmaceutical industry, waste purification and water treatment (Alwadani and Fatehi, 2018; Bhat et al., 2009; Klapiszewski et al., 2013; Naseem et al., 2016; Sen et al., 2015).

Commercial lignin is obtained from fractionation of wood fibers for use in the pulp and paper industries, so-called “black liquors”. The most widely processes used to extract lignin are sulfite, kraft, organosolv, and soda (Aro and Fatehi, 2017). Lignin is obtained by alkaline pulping in the kraft process, in which the black liquor is treated with NaOH and Na2S aqueous solution at 165–175 °C (Belgacem and Gandini, 2008; Collins et al., 2019). Lignins obtained from the sulfite pulping process are known as lignosulfonates. They are extracted using sulfurous acid and/or a sulfite salt containing magnesium, calcium, sodium, or ammonium at different pHs (Figueiredo et al., 2018).

In the kraft process, some ionizable groups as phenolic hydroxyl and sulfonic acid are introduced into the lignin structure (Klapiszewski et al., 2013). While in the production of calcium salt of lignosulfonic acid there are sulfur‐containing groups, phenolic hydroxyl groups, carboxyl groups, and Ca cations introduced in the material (Telysheva et al., 2001). Therefore, the structure of these two types of lignin is composed of a hydrophobic backbone and hydrophilic chains, which can allow the interfacial tension reduction between aqueous and oil phases (Bai et al., 2018; Sameni et al., 2018).

Some studies have demonstrated the lignin capacity to act as stabilizer of emulsions (Ago et al., 2016; Bertolo et al., 2019; Gundersen and Sjöblom, 1999; Li et al., 2016a; Qin et al., 2015; Tolosa et al., 2006). The lignin stabilization mechanism of emulsions will main depend on the size of these particles. Lignin can stabilize emulsions from adsorption on the oil / water interface if it is a macromolecule or in polymeric form, causing electrostatic and steric repulsion due to the existence of hydrophobic backbone and hydrophilic chains. However, if lignin is in colloidal or nanometric form, the main stabilizing mechanism is the Pickering-type, due to its spherical shape and great surface activity. The main properties that affect the behavior of macrolignin as an emulsifier are molecular weight, surface charge, functional groups and solubility, while for lignin nanoparticles are surface charge, size and wettability properties (Bai et al., 2018).

Several articles address researches on lignin, although a limited focus has been given to commercial initiatives for the use of lignin in the polymeric form. Furthermore, studies do not demonstrate the potential use of lignin in technological applications at a safe level (Dessbesell et al., 2020). Because of this, this study investigated the potential application of kraft lignin and calcium lignosulfonate as an emulsifier in oil-in-water emulsions. Emulsions stabilized by whey protein were also produced, aiming to compare the performance of lignins with a widely used natural emulsifier. The effect of the type and concentration of each emulsifier on the interface and emulsion characteristics were evaluated. Additionally, we also performed the in vitro cytotoxicity of different concentrations of kraft lignin and calcium lignosulfonate to assess their safety level to use in different applications.

Section snippets

Material

For the preparation of the emulsions the following ingredients were used: lignosulfonic acid calcium salt (Mw ∼18,000 g/mol -CAS Number 8061−52-7) and alkali lignin (Mw ∼10,000 g/mol; impurities 4% sulfur - CAS Number 8068−05-1) obtained from Sigma-Aldrich (St. Louis, USA) and whey protein from Arla Foods (Viby, Dinamarca). Refined sunflower oil (Bunge Alimentos S.A., Brazil) was purchased in a local market.

Emulsion preparation

Oil-in-water emulsions were prepared using the same weight ratio (10:90) in oil to

Interfacial tension and dilational rheology

The interfacial tension between sunflower oil and aqueous phases (containing LA, LG, or WPI at different concentrations) was evaluated to investigate the emulsifying capacity of each biopolymer. All the emulsifier concentrations (0.005, 0.05, and 0.5 % w/w) decreased the initial interfacial tension between water and sunflower oil (from about 25 mV/m) (Fig. 1). However, increasing the emulsifier concentration promoted a further decrease of the interfacial tension (Fig. 1). At the highest

Conclusion

The emulsifying capacity of lignin derivatives in polymeric form (kraft and calcium lignosulfonate) and whey protein were compared. The three biopolymers were able to stabilize emulsions even at very low concentrations (0.05 % w/w). However WPI-emulsions were most unstable after 28 days of storage, while lignins emulsions remained stable even with the change of pH to 7.0. At the highest lignin concentration (0.5 % w/w), emulsions with kraft lignin were more stable, because the higher sulfur

CRediT authorship contribution statement

Aline Czaikoski: Conceptualization, Faormal analysis, Investigation, Methodology, Validation, Writing - original draft. Andresa Gomes: Formal analysis, Writing - original draft. Karine Cristine Kaufmann: Formal analysis, Methodology. Raquel Bester Liszbinski: Formal analysis, Writing - original draft. Marcelo Bispo de Jesus: Conceptualization, Writing - original draft. Rosiane Lopes da Cunha: Conceptualization, Funding acquisition, Writing - original draft.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgments

This study was financed in part by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior – Brazil (CAPES) - (2952/2011) and the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) (140705/2017-1) (307168/2016-6) (156024/2018-7). The authors would also like to acknowledge the Brazilian Nanotechnology National Laboratory (LNNano) for the allocation of the equipments and the Multi-User Equipment Program grant #15/06134-4, São Paulo Research Foundation (FAPESP).

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