Optimization of Inulin Hydrolysis by Penicillium lanosocoeruleum Inulinases and Efficient Conversion Into Polyhydroxyalkanoates.

Iolanda Corrado,Giovanni Sannia,Cinzia Pezzella,Leila Birolo,Nicoletta Cascelli,Georgia Ntasi

doi:10.3389/fbioe.2021.616908

Abstract

Inulin, a polydisperse fructan found as a common storage polysaccharide in the roots of several plants, represents a renewable non-food biomass resource for the synthesis of bio-based products. Exploitation of inulin-containing feedstocks requires the integration of different processes, including inulinase production, saccharification of inulin, and microbial fermentation for the conversion of released sugars into added-value products. In this work paper, a new microbial source of inulinase, Penicillium lanosocoeruleum, was identified through the screening of a fungal library. Inulinase production using inulin as C-source was optimized, reaching up to 28 U mL–1 at the 4th day of growth. The fungal inulinase mixture (PlaI) was characterized for pH and temperature stability and activity profile, and its isoenzymes composition was investigated by proteomic strategies. Statistical optimization of inulin hydrolysis was performed using a central composite rotatable design (CCRD), by analyzing the effect of four factors. In the optimized conditions (T, 45.5°C; pH, 5.1; substrate concentration, 60 g L–1; enzyme loading, 50 U gsubstrate –1), up to 96% inulin is converted in fructose within 20 h. The integration of PlaI in a process for polyhydroxyalkanoate (PHA) production by Cupriavidus necator from inulin was tested in both separated hydrolysis and fermentation (SHF) and simultaneous saccharification and fermentation (SSF). A maximum of 3.2 g L–1 of PHB accumulation, corresponding to 82% polymer content, was achieved in the SSF. The proved efficiency in inulin hydrolysis and its effective integration into a SSF process pave the way to a profitable exploitation of the PlaI enzymatic mixture in inulin-based biorefineries.

Highlights

The biorefinery concept focuses on the sustainable conversion of renewable biomasses into a broad range of industrial products, materials, and energy
Library Screening A library of twelve fungi was assembled by choosing among strains with reported evidence in literature of inulinase production and/or for which the presence of genes belonging to the GH32 inulinase family was deduced from querying the CAZY database5 (Table 1)
The highest level of inulinase activity was detected in the extracellular media of the two abovementioned strains: about 18 U mL−1 at the 4th day and 9 U mL−1 at the 6th day, with an I/S ratio equal to 1 and 2, respectively, with these values being indicative of the prevalence of inulinase activity over the invertase one (Singh and Singh, 2010; Figure 1)

Summary

Introduction

The biorefinery concept focuses on the sustainable conversion of renewable biomasses into a broad range of industrial products, materials, and energy. PHA properties are dependent on their monomer chain length (ranging from the most common C4 monomer, butyrate, to C ≥ 6 monomers) with a composition strictly influenced by the supplied carbon source and the specific metabolic pathway activated in the cell. This variability translates into a wide range of material properties allowing this polymer to find applications in different sectors (Raza et al, 2018; Zhong et al, 2020). The use of low-cost substrates as starting feedstock for microbial fermentation represents the keystone to promote a cost-effective and sustainable exploitation of this class of biopolymers (Kumar et al, 2019; Tsang et al, 2019; Vastano et al, 2019; Guzik et al, 2020; Surendran et al, 2020)

Methods

Results

Conclusion