Optimization of welan gum extraction and purification using lysozyme and alkaline protease

Abstract

Welan gum, a natural polysaccharide produced by Sphingomonas sp. ATCC 31555, belongs to the sphingan family along with xanthan gum and gellan gum. Its structure is composed of repeating tetrasaccharide backbone, consisting of D-glucose, D-glucuronic acid, D-glucose, and L-rhamnose, with side chains containing L-rhamnose or L-mannose. Additionally, approximately half of the tetrasaccharide backbone also carry acetyl and glyceroyl groups (Kaur et al. 2014 ).This unique structural composition imparts non-gelling properties to welan gum, setting it apart from other sphingans (Wen et al. 2021 ). Furthermore, welan gum exhibits remarkable thermal stability in aqueous solutions, retaining its properties even at temperatures as high as 150 °C. It also demonstrates acid and alkali stability, maintaining viscosity under extreme pH conditions (pH 2 to 12). These outstanding characteristics have led to the extensive use of welan gum in improving building materials such as cement and concrete, as well as in applications within the household chemicals and petroleum extraction industries. In recent years, research has explored the potential application of welan gum in areas like food, cosmetics, and biodegradable materials. However, the challenges of extraction and purification, including their complexity, low efficiency, and high costs, have significantly restricted its expansion into other domains. Sphingomonas (Carmona et al. 2023 ; Baudoin et al. 2023 ; Lu et al. 2023 ; Nsengiyumva and Alexandridis 2022 ). Challenges, such as high viscosity, residual bacterial cells, carotenoids, and protein complexation, pose critical obstacles to the purification and extraction of welan gum, making the purification of the final product difficult; in particular, these challenges complicate welan gum purification and extraction in terms of impurity removal, product purification, and drying, which account for approximately 50% of the total production costs of welan gum (Kaur et al. 2014 ). Traditional extraction and purification methods involve multiple dilutions and the use of large quantities of organic solvents and chemical reagents (Roca et al. 2015 ). However, these methods result in the coprecipitation of large molecular impurities, forming irreversible aggregates. Lopes et al. subjected diluted, non-heat-treated fermentation broth to NaOH treatment followed by filtration through a 0.2 µm membrane to avoid organic reagent precipitation and welan gum coagulation caused by heating and sterilization (Lopes et al. 1994 ). However, this method showed limited effectiveness in removing proteins and carotenoid. Other promising techniques, such as size-exclusion chromatography and ion-exchange chromatography, have been partially applied in laboratory-scale preparation studies on medium-molecular-weight welan gum but have not yet been developed for industrial production mainly due to the properties and costs associated with welan gum (Zhao et al. 2022 ). In addition, the molecular weights of some low-to-medium-molecular-weight welan gums produced through cleaning with cheap substrates are close to those of proteins, making protein separation difficult. Carotenoid removal has also been a recent focus of research. For example, Zhao et al. disrupted the sorting enzyme gene (srtW) in Sphingomonas sp. ATCC 31555 strain to modify cell wall capsule morphology; they obtained welan gum with high transparency, albeit with a drastic decrease in molecular weight (Zhao et al. 2021 ). Similarly, Zhang et al. inserted the Vitreoscilla hemoglobin gene with the lacZ promoter into the carotenoid hydroxylase gene region of the ATCC 31555 chromosome to obtain an industrial strain devoid of carotenoids (Zhang et al. 2016 ). Thus, the quest for an efficient, environmentally friendly, and cost-effective extraction and purification method remains a focal point of current research.

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