Accumulation of bioplastic polyhydroxyalkanoate with different substrate forms from pretreated waste lignocellulose hydrolysate
Graphical abstract
Introduction
Rapid depletion of fossil resources and the “white pollution” caused by excessive accumulation of traditional plastic products in the environment have stimulated researchers to explore extensively bio-based and biodegradable polymers, with a view to realizing the eco-friendly and sustainable development of industrial production (Chen et al., 2009; Evangeline and Sridharan, 2019). Polyhydroxyalkanoate (PHA), which can be stored and accumulated in the cells by numerous kinds of bacteria under nutrient limited conditions, is considered as a preferred alternative to traditional petroleum-derived plastics due to their similar physical properties (Elain et al., 2016). In particular, PHA has the characteristics of biocompatibility, biodegradability and can be produced from renewable resources (Kataria et al., 2018), but it is undeniable that carbon source substrate and expensive production cost are the bottleneck of PHA industrial application. Therefore, it is urgent to seek alternative low-cost carbon substrates. Lignocellulose biomass is regarded as optimum choice of organic carbon source because of its abundance, easy availability and low cost. It is reported that the hydrolysates obtained by pretreatment from numerous agro-industrial lignocellulose wastes, such as sugarcane molasses (Lopes et al., 2014), tequila bagasse (Munoz and Riley, 2008), liquefied wood (Koller et al., 2015), rice bran (Shamala et al., 2012), maple wood (Pan et al., 2012), barley (Bhatia et al., 2018), wheat straw (Cesario et al., 2014) have been utilized for PHA production. However, the above research basically adopts pure bacteria culture, whose cost is relatively high.
Mixed microbial culture (MMC) in activated sludge for PHA production, possessing the advantages of easy process control, free selection of microorganisms and long-term stable operation of the system, has been proposed as an alternative process to high-cost pure culture fermentation (Tu et al., 2019; Valentino et al., 2017). However, the substrates such as papermaking wastewater, olive oil wastewater, mixed food wastewater and urban wastewater contain a lot of carbohydrates, which are often used to accumulate glycogen rather than PHA when used as carbon source (Amulya et al., 2015; Dai and McDonald, 2014; Jiang et al., 2012; Morgan-Sagastume, 2016). Volatile fatty acids (VFAs), as an organic fermentation product, are one of the most fascinating substrate for PHA production. Generally speaking, it is the most appropriate choice to ferment waste carbon sources into intermediate product VFAs prior to the culture selection and PHA accumulation. VFAs obtained from co-fermentation of pretreated waste lignocellulose hydrolysate and sewage has been tested as carbon source for the production of poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) from numerous bacterial systems studied by our group (Li et al., 2020). Although the production of PHA was improved, the proportion of 3-hydroxyvalerate (HV) monomer was only 16 %. Some studies have shown that the introduction of flexible monomers (such as HV) into poly (3-hydroxybutyrate) (PHB) to form copolymers can improve its brittleness and make it have excellent physical and thermal properties, which remedy the defects that 3-hydroxybutyrate (HB) and HV have better properties than themselves (Cal et al., 2021). The use of co-substrate as feedstock for PHA production has been regarded as the most promising and economically feasible method to increase PHA production and HV monomer ratio (Dalal et al., 2013; Lemechko et al., 2019; Yin et al., 2020).
Therefore, the purpose of this study was to explore the feasibility of using pretreated waste lignocellulose hydrolysate and its fermented product VFAs (mainly acetic acid, propionic acid and butyric acid) as co-substrates, and MMC derived from activated sludge instead of pure bacteria culture to produce PHA. The content of PHA synthesized from single VFAs, lignocellulose hydrolysate and VFAs co-substrate two carbon sources under different carbon source concentration and C/N ratio was monitored. Further, the structural components of extracted PHA were systematically characterized by FT-IR, NMR (1H & 13C) and GC–MS, and molecular weight, thermal properties of PHA synthesized by two carbon sources were compared. The extracted PHA was further purified to prepare the film, and its mechanical properties were investigated.
Section snippets
Materials
The lignocellulose hydrolysate was prepared in a high temperature and high pressure reactor with dried poplar wood flour (particle size 40–80 meshes) as previously described (Yin et al., 2019). VFAs were produced via co-fermentation of activated sludge as the source of mixed bacteria and lignocellulose hydrolysate as carbon source under the conditions of 35 ± 1 ℃, pH value of 8 ± 0.2 and C/N ratio of 60 ± 2. The detailed fermentation process was as previously described by Li et al. (2019a). The
Effects of carbon source types on PHA production
PHA accumulation experiments were conducted for two carbon sources (single VFAs, lignocellulose hydrolysate and VFAs co-substrate) to evaluate the variation of COD and PHA content over time. The obtained hydrolysate contained reducing sugar of 530.3 mg g−1, furfural of 512.6 mg L−1 and 5-HMF of 239.3 mg L−1, respectively. From Fig. 1a and b, the uptake rate of single VFAs carbon source by microbes was significantly higher than that of lignocellulose hydrolysate and VFAs co-substrate. The former
Conclusions
In this study, the rule of PHA accumulation and the characterization from single VFAs, lignocellulose hydrolysate and VFAs co-substrate two carbon sources were investigated. Compared with single VFAs carbon source, the addition of co-substrate increased the PHA production to 3049 mg COD/L (1.73 mg mL−1, 0.636 g PHA/g VSS), especially the proportion of HV monomer reached 44.9 % at C/N ratio of 33. For the two carbon sources, limited nitrogen source can promote the accumulation of PHA.
CRediT authorship contribution statement
Dongna Li: Methodology, Investigation, Data acquisition, Data analysis, Manuscript writing and revising. Jianing Li: Data analysis, Manuscript Writing - Review & Editing. Xiaojun Ma: Conceptualization, Methodology, Supervision, Data analysis, Manuscript Writing - Review & Editing.
Declaration of Competing Interest
The authors declare that there is no conflict of interests regarding the publication of this article.
Acknowledgements
The authors are grateful for the financial supports from Science Foundation of Tianjin Municipal Education Commission (2019ZD039), Tianjin Natural Science Foundation (18JCYBJC90100), and Opening Project Fund of Key Laboratory of Biology and Genetic Resources of Rubber Tree, Ministry of Agriculture and Rural Affairs, P. R. China/ State Key Laboratory Breeding Base of Cultivation & Physiology for Tropical Crops/Danzhou Investigation & Experiment Station of Tropical Crops, Ministry of Agriculture
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